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Team:TU Munich/Project/Limonene
Larakuntz (Talk | contribs)
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[[File:Gruppe_limonen_TUM12.jpg|200px|thumb||Responsible: Lara Kuntz and Andrea Richter]]
'''A common saying is "If life gives you lemons - make lemonade". As of iGEM it can only be "If life gives you limonene - make beer"'''.
'''A Common Saying Is "If Life Gives You Lemons - Make Lemonade". For Us It Is Rather "If Life Gives You Limonene - Make Beer"'''.
The obvious thing to do, isn't it? Beer with lemonade is a very popular beverage throughout Germany, e.g. "Radler", "Alsterwasser", "Russ'n". So why not take the shortcut and skip the intermediary?
Just think of the refreshing sensation of lemons paired with the complex richness of a <s>chilled</s> ''tepid'' brew.
'''Limonene''' is a cyclic terpene and a major constituent of several citrus oils. D-Limonene is used as a component of flavorings and fragrances since it has an orange/lemon-like odor. Limonene has been shown to inhibit rat mammary and other tumor development [[http://www.ncbi.nlm.nih.gov/pubmed?term=Tsuda%20limonene%202004 Tsuda et al., 2004]]. Being an excellent solvent of cholesterol, d-limonene also has been used clinically to dissolve cholesterol-containing gallstones. Because of its gastric acid neutralizing effect and its support of normal peristalsis, it has also been used for relief of heartburn [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
D-limonene is used as a component of flavorings and fragrances since it has an orange/lemon-like odor. '''Limonene''' has been shown to inhibit rat mammary and other tumor development [[http://www.ncbi.nlm.nih.gov/pubmed?term=Tsuda%20limonene%202004 Tsuda et al., 2004]]. Being an excellent solvent of cholesterol, d-limonene also has been used clinically to dissolve cholesterol-containing gallstones. Because of its gastric acid neutralizing effect and its support of normal peristalsis, it has also been used for relief of heartburn [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
Producing the flavoring substance limonene in our beer might result in a fresh, '''lemon-like taste''' on the one hand. On the other hand, we might have '''beneficial effects on health''' such as preventive activity against cancer, dissolution of gallstones and relief of heartburn.
== Background and principles ==
We have achieved our goal of a beer with limonene. We have successfully cloned (+)-limonene synthase 1 into our newly established yeast expression vector pTUM100. We transformed ''Saccharomyces cerevisiae'' with the plasmid carrying limonene synthase. We '''proved expression''' of limonene synthase via western blot. We verified the '''functionality of (+)-limonene synthase 1''' by in vitro assay with purified limonene synthase on the one hand. On the other hand, we proved functional '''limonene production in yeast cell culture''' via headspace GC-MS. Furthermore, we analyzed differences in protein expression in yeast depending on existence of the yeast '''consensus sequence'''. To survey the toxic concentration of limonene for yeast cells, a '''toxicity assay''' was performed. We showed an inhibition of growth at 1 mM and a lethal effect at 100 mM. These concentrations should not be reached by expression of limonene synthase in yeast.
Last but not least, we have been able to proof the '''production of limonene in the beers''' we brewed. Hence, we have brewed''' iGEM's first SynBio beer''' containing limonene.
Limonene is a cyclic terpene and a major constituent of several citrus oils (orange, lemon, mandarin, lime and grapefruit). It is a chiral liquid with the molecular mass of 136.24 g/mol. The (R)-enantiomer smells like oranges and is content of many fruits, while the (S)-enantionmer has a piney odor [[http://www.ncbi.nlm.nih.gov/pubmed/11605760 Fietzek et al., 2001]]. Therefore D-Limonene ((+)-Limonene, (R)-enantiomer) is used as a component of flavorings and fragrances.
Limonene is a cyclic terpene and a major constituent of several citrus oils (orange, lemon, mandarin, lime and grapefruit). It is a chiral liquid with the molecular mass of 136.24 g/mol. The (R)-enantiomer smells like oranges and is content of many fruits, while the (S)-enantionmer has a piney odor [[http://www.ncbi.nlm.nih.gov/pubmed/11605760 Fietzek et al., 2001]]. Therefore, d-limonene ((+)-limonene, (R)-enantiomer) is used as a component of flavorings and fragrances.
=== Biosynthesis ===
Limonene is produced by '''limonene synthase''' which uses geranyl pyrophosphate (GPP) as educt which is the universal precursor of monoterpenoids. (+)-limonene synthase from ''Citrus limon'' consists of 606 aminoacids (EC=4.2.3.20) and catalyzes the following reaction: '''Geranyl pyrophosphate = (+)-(4R)-limonene + diphosphate'''.
Limonene is produced by '''limonene synthase'''. Limonene synthase uses geranyl pyrophosphate (GPP), which is the universal precursor of monoterpenoids, as educt. (+)-limonene synthase from ''Citrus limon'' consists of 606 aminoacids (EC=4.2.3.20) and catalyzes the following reaction: '''Geranyl pyrophosphate = (+)-(4R)-limonene + diphosphate''' (see Fig. 1).
[[File:TUM12_ReactionLimoneneSynthase.jpg|thumb|900px|Fig. 1: Reaction catalyzed by limonene synthase.]]
''Saccharomyces cerevisiae'' produces geranyl pyrophosphate via the mevalonate pathway where it occurs exclusively as an intermediate of farnesyl pyrophosphate (FPP) synthesis [[http://www.ncbi.nlm.nih.gov/pubmed/17096665 Oswald et al., 2007]]. It has been established that S. cerevisiae has enough free GPP to be used by exogenous monoterpene synthases to produce monoterpenes under laboratory and vinification conditions [[http://www.ncbi.nlm.nih.gov/pubmed/18155949 Herrero et al., 2008]][[http://www.ncbi.nlm.nih.gov/pubmed/17096665 Oswald et al., 2007]].
''Saccharomyces cerevisiae'' produces geranyl pyrophosphate via the mevalonate pathway (see Fig. 2). GPP occurs as an intermediate of farnesyl pyrophosphate (FPP) synthesis [[http://www.ncbi.nlm.nih.gov/pubmed/17096665 Oswald et al., 2007]]. It has been established that ''S. cerevisiae'' has enough free GPP to be used by exogenous monoterpene synthases to produce monoterpenes under laboratory and vinification conditions [[http://www.ncbi.nlm.nih.gov/pubmed/18155949 Herrero et al., 2008], [http://www.ncbi.nlm.nih.gov/pubmed/17096665 Oswald et al., 2007]].
=== The molecular and physiological effects of limonene ===
[[File:TUM12_TerpeneSynthesis.png|thumb|900px|Fig. 2: Simplified isoprenoid pathway in ''S. cerevisiae'', including the branch point to linalool. Abbreviations: HMG-CoA, 3-hydroxy-3-methylglutaryl coenzyme A; IPP, isopentenyl pyrophosphate; GPP, geranyl pyrophosphate; FPP, farnesyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; HMGR, HMG-CoA reductase; FPPS, FPP synthase; LIS, linalool synthase. [<html><a href="http://www.ncbi.nlm.nih.gov/pubmed/20675444">Rico et al., 2010</a></html>]]]
[[File:TUM12_TerpeneSynthesis.png|thumb|900px|Fig. 2: Simplified isoprenoid pathway in S. cerevisiae, including the branch point to linalool. Abbreviations: HMG-CoA, 3-hydroxy-3-methylglutaryl coenzyme A; IPP, isopentenyl pyrophosphate; GPP, geranyl pyrophosphate; FPP, farnesyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; HMGR, HMG-CoA reductase; FPPS, FPP synthase; LIS, linalool synthase. [<html><a href="http://www.ncbi.nlm.nih.gov/pubmed/20675444">Rico et al., 2010</a></html>]]]
====Flavour and Aliment====
=== Molecular and Physiological Effects of Limonene ===
Because of its pleasant Citrus flavour and very low toxicity (oral LD50 for mice = 5.6 and 6.6 g/kg body weight), D-Limonene is widely used as a flavor and fragrance additive. Therefore it is listed in the Code of Federal Regulations as generally recognized as safe ('''GRAS''') [[http://www.fda.gov/downloads/Food/FoodIngredientsPackaging/GenerallyRecognizedasSafeGRAS/GRASListings/UCM264589.pdf FDA]] for a flavoring agent and can be found in common food items such as fruit juices, soft drinks, baked goods, ice cream, and pudding in typical concentrations of 50 ppm till 2,500 ppm, respectively [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]]. Hence the normal daily consume of D-Limonene is 0,27 mg/kg body weight per day [[http://www.mri.bund.de/fileadmin/Institute/PBE/Sekundaere_Pflanzenstoffe/Monoterpene.pdf Watzl, 2002]]. As '''natural compound''' of Plants Limonene has practical advantages with regard to availability, suitability for oral apllication, regulatory approval and mechanisms of action and does not pose a mutagenic, carcinogenic, or nephrotoxic risk to humans [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
====Cancer inhibition====
====Flavor and Aliment====
Monoterpenes have anti carcinogenic effects in animal experiments. It has been shown to inhibit rat mammary, gastric, lung and skin tumor development by several discussed mechanisms like apoptosis induction and modulation of oncogene signal transduction [[http://www.ncbi.nlm.nih.gov/pubmed?term=Tsuda%20limonene%202004 Tsuda et al., 2004]] [[http://www.mri.bund.de/fileadmin/Institute/PBE/Sekundaere_Pflanzenstoffe/Monoterpene.pdf Watzl, 2002]]. So D-limonene induces phase I and phase II carcinogen-metabolizing enzymes (cytochrome p450), which metabolize carcinogens to less toxic forms and prevent the interaction of chemical carcinogens with DNA. It also inhibits tumor cell proliferation, acceleration of the rate of tumor cell death and/or induction of tumor cell differentiation. Furthermore, D-Limonene regulate cell growth and/or transformation by inhibiting protein isoprenylation [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
Because of its pleasant citrus flavour and very low toxicity (oral LD50 for mice = 5.6 and 6.6 g/kg body weight), d-limonene is widely used as a flavor and fragrance additive. It is listed in the Code of Federal Regulations as a generally recognized as safe ('''GRAS''') flavoring agent [[http://www.fda.gov/downloads/Food/FoodIngredientsPackaging/GenerallyRecognizedasSafeGRAS/GRASListings/UCM264589.pdf FDA]]. It can be found in common food items such as fruit juices, soft drinks, baked goods, ice cream, and pudding in typical concentrations of 50 ppm to 2,500 ppm [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]]. The normal daily consume of d-limonene is 0.27 mg/kg body weight per day [[http://www.mri.bund.de/fileadmin/Institute/PBE/Sekundaere_Pflanzenstoffe/Monoterpene.pdf Watzl, 2002]]. As '''natural compound''' of plants, limonene has practical advantages with regard to availability, suitability for oral application, regulatory approval and mechanisms of action. It does not pose a mutagenic, carcinogenic or nephrotoxic risk to humans [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
====Solvent for Gallstones====
Furthermore it is used as excellent '''solvent of cholesterol''', therefore d-limonene has been used clinically to dissolve cholesterol-containing gallstones [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]]. A study with 200 patients reported a direct infusion of 20-30 ml D-Limonene (97% solution) completely or partially dissolved gallstones in 141 patients. Stones completely dissolved in 96 cases (48%); partial dissolution was observed in 29 cases (14.5%); and in 16 cases (8%) complete dissolution was achieved with the inclusion of hexamethaphosphate (HMP), a chelating agent that can dissolve bilirubin calcium stones [[http://www.ncbi.nlm.nih.gov/pubmed/1988264 Igimi et al., 1991]]. Because of its gastric acid neutralizing effect and its support of normal peristalsis, it also has been used for relief of heartburn [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
Monoterpenes show anticarcinogenic effects in animal experiments. They have been shown to inhibit rat mammary, gastric, lung and skin tumor development by several discussed mechanisms such as apoptosis induction and modulation of oncogene signal transduction [[http://www.ncbi.nlm.nih.gov/pubmed?term=Tsuda%20limonene%202004 Tsuda et al., 2004], [http://www.mri.bund.de/fileadmin/Institute/PBE/Sekundaere_Pflanzenstoffe/Monoterpene.pdf Watzl, 2002]]. d-limonene induces phase I and phase II carcinogen-metabolizing enzymes such as cytochrome p450. These metabolize carcinogens into less toxic forms on the one hand. On the other hand, they prevent the interaction of chemical carcinogens with DNA. Limonene has also been shown to inhibit tumor cell proliferation, to accelerate the rate of tumor cell death and induce tumor cell differentiation. Furthermore, d-limonene regulates cell growth and/or transformation by inhibiting protein isoprenylation [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
====Solvation of Gallstones====
d-limonene is used as excellent '''solvent of cholesterol''', therefore it has been used clinically to dissolve cholesterol-containing gallstones [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]]. A study with 200 patients showed that direct infusion of 20-30 ml d-limonene (97% solution) completely or partially dissolved gallstones in 141 patients. Gallstones completely dissolved in 96 cases (48%); partial dissolution was observed in 29 cases (14.5%); and in 16 cases (8%) complete dissolution was achieved with the inclusion of hexamethaphosphate (HMP), a chelating agent that can dissolve bilirubin calcium stones [[http://www.ncbi.nlm.nih.gov/pubmed/1988264 Igimi et al., 1991]]. Because of its gastric acid neutralizing effect and its support of normal peristalsis, it has also been used for relief of heartburn [[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]].
===BioBricks===
<div class="mfull bezel">
==== [http://partsregistry.org/Part:BBa_K801060 BBa_K801060] '''(+)-Limonene synthase 1''' with '''Strep-tag''' and yeast '''consensus sequence''' ====
==== [http://partsregistry.org/Part:BBa_K801060 BBa_K801060] '''(+)-Limonene synthase 1''' with '''''Strep''-tag''' and yeast '''consensus sequence''' ====
This part contains the (+)-Limonene Synthase 1 of Citrus limon. It is preceeded by the yeast consensus sequence for improved expression and carries a C-Terminal Strep-Tag for purification or detection by westernblot. It is an improved version of BBa_I742111.
This part contains (+)-limonene synthase 1 of ''Citrus limon''. It is preceeded by the yeast consensus sequence for improved expression and carries a C-Terminal ''Strep''-Tag for purification or detection by westernblot. It is an improved version of BBa_I742111.
RFC 25 compatible
==== [http://partsregistry.org/Part:BBa_K801061 BBa_K801061] '''(+)-Limonene synthase 1''' coding region from ''Citrus limon'' ====
Improved version of BBa_I742111.
Does not contain stop codon. Needs to be used with RFC 25.
Does not contain a stop codon. Needs to be used with RFC 25.
'''Further information:'''
==== [http://partsregistry.org/Part:BBa_K801062 BBa_K801062] '''(+)-limonene synthase 1 expression cassette for yeast'''====
This part can be used to express ''Citrus limon'' (+)-limonene synthase 1 in yeast. The expression is controlled by TEF1 promoter and CYC1 terminator.
==== [http://partsregistry.org/Part:BBa_K801063 BBa_K801063] '''(+)-limonene synthase 1 expression cassette for yeast''' ====
This part can be used to express ''Citrus limon'' (+)-limonene synthase 1 in yeast. The expression is controlled by yeast TEF1 promoter and yeast TEF1 terminator.
This part can be used to express ''Citrus limon'' (+)-limonene synthase 1 in yeast. The expression is controlled by yeast TEF2 promoter and yeast CYC1 terminator.
==== [http://partsregistry.org/Part:BBa_K801065 BBa_K801065] '''(+)-Limonene synthase 1''' with '''Strep-Tag''' ====
==== [http://partsregistry.org/Part:BBa_K801065 BBa_K801065] '''(+)-Limonene synthase 1''' with '''''Strep''-Tag''' ====
This part contains the coding region of (+)-limonene synthase from Citrus limon with a C-terminal Strep-tag. This part is based on BBa_K801061.
This part contains the coding region of (+)-limonene synthase from Citrus limon with a C-terminal ''Strep''-tag. This part is based on BBa_K801061.
==== [http://partsregistry.org/Part:BBa_K801066 BBa_K801066] '''(+)-Limonene synthase 1''' with yeast '''consensus sequence''' for improved expression====
This part is second version of BBa_K801060. It contains (+)-limonene synthase and the consensus sequence for enhanced expression in yeast. It does not contain a C-terminal Strep tag as BBa_K801060. The sequence does not contain a stop codon so that RFC 25 has to be used.
This part is second version of BBa_K801060. It contains (+)-limonene synthase and the consensus sequence for enhanced expression in yeast. It does not contain a C-terminal ''Strep''-tag as BBa_K801060. The sequence does not contain a stop codon so that RFC 25 has to be used.
==== Gel Picture of finished construct ====
==== Gel Picture of Finished Constructs ====
[[file:TUM12_limonenegel.png|thumb|300px|right|Fig. 3: Gel electrophoresis of [http://partsregistry.org/Part:BBa_K801060 K801060] and [http://partsregistry.org/Part:BBa_K801061 K801061] after analytical restrigtion digest with EcoR1 and Pst1.]]
(+)-Limonene synthase 1 coding region without yeast consensus sequence [[http://partsregistry.org/Part:BBa_K801061 BBa_K801061]] in pSB1C3 is shown next to (+)-limonene synthase 1 with ''Strep''-tag and yeast consensus sequence [[http://partsregistry.org/Part:BBa_K801060 BBa_K801060]] after restriction digest with EcoR1 and Pst1 restriction enzymes. To check success of ligation, DNA fragments were separated by agarose gel-electrophoresis using ethidium bromide as a nucleic acid stain. As expected, the 1665 bp fragment of [[http://partsregistry.org/Part:BBa_K801061 BBa_K801061]] and the 1708 bp fragment of [[http://partsregistry.org/Part:BBa_K801060 BBa_K801060]] were detected additionally to the pSB1C3 vector that has 2070 bp (see Fig. 3).
To compare limonene synthase expression in yeast depending on yeast consensus sequence, we produced duplicates of biobricks; one of each has the consensus sequence, the other one does not. For yeast expression experiments, the biobricks were cloned into the yeast expression vector (pTUM100) recently designed by us.
==== Investigation of the yeast consensus sequence ====
==== Investigation of Yeast Consensus Sequence ====
[[file:TUM12_consensus.png|thumb|450px|right|Fig. 4: Comparison of limonene synthase biobricks ([http://partsregistry.org/Part:BBa_K801065 BBa_K801065] and [http://partsregistry.org/Part:BBa_K801060 BBa_K801060]) with and without yeast consensus sequence.]]
[[http://www.ncbi.nlm.nih.gov/pmc/articles/PMC340751/ Hamilton et al., 1987]] reported of a consensus sequence upstream of the AUG start codon in yeast.
Although not as strong as the mammalian Kozak translation initiation sequence, the yeast consensus sequence is thought to have a 2–3-fold effect on the efficiency of translation initiation [[http://tools.invitrogen.com/content/sfs/manuals/pyes2_man.pdf pYES2 manual]].
We designed duplicates of limonene synthase encoding biobricks; one having the yeast consensus sequence, the other one not having the consensus sequence.
We have only been able to show slight differences in expression between the two biobricks [[http://partsregistry.org/Part:BBa_K801065 BBa_K801065] and [http://partsregistry.org/Part:BBa_K801060 BBa_K801060]] via coomassie staining and western blot. The difficulties of showing the difference via SDS-page may result from variations in the amount of protein applied.
Our in vivo analysis strongly indicates that the consensus sequence does lead to a 2-3-fold enhanced expression in yeast, though (see Fig. 8B). This is consistent with findings of others [[http://tools.invitrogen.com/content/sfs/manuals/pyes2_man.pdf pYES2 manual]].
==== In vitro detection of limonene====
====Purification of Recombinant Limonene Synthase====
[[File:TUM12_SApurification.png|thumb|200px|left|Fig. 5: Streptavidin Affinity (SA) chromatography.]]
[[File:TUM12_gelfiltration.png|thumb|450px|right|Fig. 6: Analytical gelfiltration of limonene synthase.]]
'''Streptavidin affinity chromatography'''<br>
The yeast cell extract that was obtained by cell lysis using glass beads of 0.5 mm was centrifuged at 11000 RPM in an SLA-3000 rotor for 60 minutes and subsequently dialysed against 5 liters of 1x Streptavidin Affinity buffer (SA-buffer) over night. The cell extract was then filtrated using a syringe filter with a pore size of 0.45μm and susequently applied on an SA-column. After sample application, the column was washed with 1x SA buffer until a base line was reached. Subsequently, bound protein was eluted using 5 mM Biotin in 1x SA buffer. The chromatogram of the purification is shown on the left side.<br><br>
[[file:TUM12_limonene_invivo.png|450px|thumb|right|Fig. 5: Spectrum of in vitro detection of limonene (enzyme assay with [http://partsregistry.org/wiki/index.php?title=Part:BBa_K801060 BBa_K801060]) and reference spectrum.]]
'''Gel Filtration of Purified Protein'''
The protein sample obtained was concentrated using a centrifugation concentrator with a molecular size limit of 30 kDa subsequent filtration. 250μl of the solution were applied to an analytical gel filtration column Superdex 200 10/30 with 1x PBS as running buffer at a flow rate of 0.5 ml/min. In the chromatogram (shown in the figure on the right in section B), there is an aggregate peak at the exclusion limit that may be caused by the preceding concentration and a major peak at an elution volume of 13.580 ml. The calibration line that was obtained from the calibration proteins b-amylase, alcohol dehydrogenase, BSA, ovalbumine, carboanhydrase, cytochrome C and aprotinin filtrated with the same experimental setup resulted in a regression line with the formula y = -39206 x + 3.3463. Using this formula and the elution volume of the limonene synthase, an apparent molecular mass of 70.1 kDa could be determined for the produced limonene synthase. This fits quite well the theoretical molecular mass that was calculated using [http://web.expasy.org/cgi-bin/protparam/protparam ExPASy ProtParam] to be 65977.1 Da. The four kilo daltons difference could be caused by posttranslational modifications. This hypothesis should be tested using mass spectrometry.
To test the functionality of purified limonene synthase in vitro, we used an optimized protocol of an enzyme assay with extraction of limonene [[http://www.ncbi.nlm.nih.gov/pubmed/17662687 Landmann et al, 2007]]. The limonene synthase was purified via ''Strep''-tag. The enzyme assay was carried out in 25 mM Tris-HCl buffer with 5% Glycerol, 1 mM DTT and cofactors (10 mM MgCl2, 1 mg/ml BSA). 50 µM substrate (geranyl pyrophosphate) and 10 ng purified recombinant limonene synthase were added to the reaction batch. Negative controls were reaction batches without enzyme. The reaction was incubated for 15 min at room temperature. Afterwards, limonene was extracted with pentane, dried with sodiumsulfate and reduced under a stream of nitrogen. Three replicates were done of both sample and negative control.
To test functionality of purified Limonenesynthase in vitro we used an optimized protocol of an enzyme assay with previous extraction of [[http://www.ncbi.nlm.nih.gov/pubmed/17662687 Landmann et al, 2007]]. The enzyme assay was carried out in 25 mM Tris-HCl buffer with 5% Glycerol, 1mM DTT and cofactors (10 mM MgCl2, 1 mg/ml BSA). An identified amount of substrate (geranyl pyrophosphate) and purified recombinant limonene synthase. The mixture was extracted with pentane, dried with sodiumsulfate and reduced under a stream of nitrogen.
The pentane extracts were analyzed with gas chromatography-mass spectrometry ("5890 Series II GC" coupled to a "Finnigan Mat 55 S MS") to identify the enzymatically synthesized products.
All enzyme reactions (three replicates) led to the production of limonene while the negative controls did not show limonene. Therefore, we showed that our purified '''limonene synthase is functional and leads to the production of limonene'''.
==== In vivo detection of limonene====
[[file:TUM12_limoneneinvivo.png|450px|thumb|right|Fig. 6: Detection of limonene in headspace above cell culture supernatant. [A] Spectrum of limonene obtained when analyzing cell culture that was transformed with pTUM104 containing construct of [http://partsregistry.org/wiki/index.php?title=Part:BBa_K801060 BBa_K801060]. [B] Overview about different measurements.]]
Because limonene is volatile, we expected an arbitrarily amount of limonene in the gaseous phase above the cell culture supernatant as in the cell culture. Therefore, we measured limonene via headspace (SPME needle) GC-MS.
Because limonene is a VOC (volatile organic compound) [[http://www.ncbi.nlm.nih.gov/pubmed/15763095 Pierucci et al., 2005]], we expected limonene to be present in the gaseous phase above the cell culture as well as in the cell culture. Therefore, we measured limonene via headspace (SPME needle) GC-MS.
We showed limonene to be produced by yeast that was transformed with pTUM104 carrying limonene synthase coding regions (see [http://partsregistry.org/wiki/index.php?title=Part:BBa_K801061 BBa_K801061] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K801060 BBa_K801060]).
We showed limonene to be produced by the yeasts that were transformed with pTUM104 carrying limonene synthase coding regions (see [http://partsregistry.org/wiki/index.php?title=Part:BBa_K801061 BBa_K801061] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K801060 BBa_K801060]).
We detected a greater amount of limonene in the sample that contained limonene synthase with consensus sequence. Hence, we showed that the yeast consensus sequence might increase the expression of limonene synthase and therefore might lead to enhanced limonene production.
We detected more limonene in the sample that contained a limonene synthase with consensus sequence. Hence, we showed that the yeast consensus sequence might increase the expression of limonene synthase and therefore lead to enhanced limonene production.
Furthermore, we were not able to detect a significant difference between samples that had additional GPP (educt) versus the ones that did not. This might be due to the inability of GPP to diffuse into the cells (hydrophilic character). Since we were able to detect limonene in both samples, it implies that the GPP present in the cells is sufficient for limonene production. This is consistent with the findings of [http://www.ncbi.nlm.nih.gov/pubmed/18155949| [Herrero et al., 2008]] that showed that ''S. cerevisiae'' cells (from laboratory and wine strains) contain enough free GPP to be catalytically transformed by monoterpene synthases into monoterpenes.
Furthermore, we could not detect a significant difference between samples that had additional GPP (educt) versus the ones that did not. This might be due to the inability of GPP to diffuse into the cells (hydrophilic character). Since we were able to detect limonene in both samples, it implies that the GPP present in the cells is sufficient for limonene production. This is consistent with the findings of [http://www.ncbi.nlm.nih.gov/pubmed/18155949| [Herrero et al., 2008]] that showed that S. cerevisiae cells (from laboratory and wine strains) contain enough free GPP to be catalytically transformed by monoterpene synthases into monoterpenes.
==== Detection of limonene in beer====
[[File:TUM12_SPME.jpg|200px|thumb|left|Fig. 7: Preparation of sample for GC-MS with SPME.]]
[[File:TUM12_LSbeer.jpg|200px|thumb|right||Bildbeschriftung]]
[[File:TUM12_LSbeer.jpg|200px|thumb|right||Fig. 10: iGEM's first and finest SynBio Beer with limonene.]]
A first attempt to use our genetically engineered yeasts to brew a SynBio Beer were conducted using a transient transfection with a constitutive promoter. The drawback is that in the gyle the selection pressure is not preserved and the loss of the plasmid is possible.<br>
A first attempt to use our genetically engineered yeasts to brew a SynBio Beer were conducted using a transient transfection with a constitutive promoter. A drawback might be that the selection pressure might not be preserved in the gyle and hence the loss of the plasmid might be possible. Therefore, we also performed brewing experiments with yeast that carried genome integrated limonene synthase.<br>
Three liters of gyle were inoculated with 100ml of a stationary yeast culture grown in YPD that was transiently transfected with a plasmid harboring a constitutive expression cassette for the limonene synthase.
Three liters of gyle were inoculated with 100ml of a stationary yeast culture grown in YPD.
We analyzed this first beer for limonene content via headspace (SPME needle) GC-MS. Unfortunately we could not yet proof a significant difference between the beer containing limonene and the negative control beer. This might be due to a loss of the plasmid which encodes limonene synthase. We will try to integrate the limonene synthase expression cassette into the genome of yeast and afterwards we will repeat the experiment.
On the one hand, we have been able to brew a beer with a yeast strain that was transformed with a vector carrying a constitutive expression cassette for limonene. On the other hand, we brewed a beer with a yeast strain that carried limonene synthase after genome integration. We analyzed the beers for limonene content via headspace (SPME needle) GC-MS. We have been able to detect limonene to be produced in both beers.
[[file:TUM12_limonene_in_beer.png|thumb|center|600px| Fig. 11: GC spectra obtained from GC-MS analysis of brewed beer. Brown: Beer brewed with yeast that carries pTUM101 with limonene synthase. Green: Beer brewed with yeast that supposedly carries limonene synthase in genome (after genome integration).]]
To establish whether limonene has an effect on yeast cells , we inoculated three different yeast strains with different concentrations of limonene. Limonene was added to the medium and the used yeast strains were the laboratory strain INVSc1, a strain which is used for brewing beer and a strain which can be purchased in a supermarket.
To establish whether limonene has an effect on yeast cells, we inoculated three different yeast strains with different concentrations of limonene. Limonene was added to the medium. We examined the laboratory strain INVSc1, a strain which is used for brewing beer and a strain which can be purchased in grocery stores.
Limonene at high concentrations affects the growth of yeast cells. We could show an inhibition of growth at 1 mM and even a lethal effect at 100 mM. At lower concentrations (1 µM, 10 µM, 100 µM) no inhibition could be observed. The growth rates of yeast cells which were incubated with low concentrations of limonene do not show a difference compared to the negative control (incubation of analogous yeast strains with YPD without limonene).
The in vivo GCMS detection of limonene [B] displayed a concentration of 50 µM. Hence the amount of limonene we will produce with the modified yeast will not reach a toxic concentration at all.
[[File:TUM12_Toxicity_Limonene.png|800px|thumb|center|Evaluation of the Toxicity Assay for Limonene.]]
At high concentrations, limonene affects the growth of yeast cells. We could show an inhibition of growth at 1 mM and even a lethal effect at 100 mM. At lower concentrations (1 µM, 10 µM, 100 µM) no inhibition could be observed. The growth rates of yeast cells which were incubated with low concentrations of limonene do not show a difference compared to the negative control (incubation of analogous yeast strains with YPD without limonene).
[[File:TUM12_Toxicity_Limonene.png|800px|thumb|center|Fig. 12: Limonene toxicity assay evaluation.]]
*[[http://www.ncbi.nlm.nih.gov/pubmed/11605760 Fietzek et al., 2001]] Fietzek, C., Hermle, T., Rosenstiel, W., Schurig, V. (2001) Chiral discrimination of limonene by use of beta-cyclodextrin-coated quartz-crystal-microbalances (QCMs) and data evaluation by artificial neuronal networks. ''Fresenius J Anal Chem.'', 371(1):58-63.
*[[http://www.ncbi.nlm.nih.gov/pmc/articles/PMC340751/ Hamilton et al., 1987]] Hamilton, R., Watanabe, C. K., De Boer, A. H. (1987) Compilation and comparison of the sequence context around the AUG startcodons in ''Saccharomyces cerevisiae'' mRNAs. ''Nucleic Acids Res.'', 15(8):3581–3593.
*[[http://www.ncbi.nlm.nih.gov/pubmed/18155949 Herrero et al., 2008]] Herrero, O., Ram ́on, D., and Orejas, M. (2008). Engineering the ''Saccharomyces cerevisiae'' isoprenoid pathway for de novo production of aromatic monoterpenes in wine. ''Metab Eng'', 10(2):78–86.
*[[http://www.ncbi.nlm.nih.gov/pubmed/1988264 Igimi et al., 1991]] Igimi, H., Tamura, R., Toraishi, K., Yamamoto, F., Kataoka, A., Ikejiri, Y., Hisatsugu, T., Shimura, H. (1991). Medical dissolution of gallstones. Clinical experience of d-limonene as a simple, safe, and effective solvent. '''Dig Dis Sci.''', 36(2):200-8.
*[[http://www.ncbi.nlm.nih.gov/pubmed/12084056 Lücker et al., 2002]] Lücker, J., El Tamer, M. K., Schwab, W., Verstappen, F. W. A., van der Plas, L. H. W., Bouwmeester, H. J., and Verhoeven, H. A. (2002). Monoterpene biosynthesis in lemon (''Citrus limon''). cDNA isolation and functional analysis of four monoterpene synthases. ''Eur J Biochem'', 269(13):3160–71.
*[[http://www.ncbi.nlm.nih.gov/pubmed/17096665 Oswald et al., 2007]] Oswald, M., Fischer, M., Dirninger, N., and Karst, F. (2007). Monoterpenoid biosynthesis in ''Saccharomyces cerevisiae''. ''FEMS Yeast Res'', 7(3):413–21.
*[[http://www.ncbi.nlm.nih.gov/pubmed/15763095 Pierucci et al., 2005]] Pierucc, P., Porazzi, E., Martinez, MP., Adani, F., Carati, C., Rubino, FM., Colombi, A., Calcaterra, E., Benfenati, E. (2005). Volatile organic compounds produced during the aerobic biological processing of municipal solid waste in a pilot plant.''' Chemosphere''', 59(3):423-30.
*[[http://www.ncbi.nlm.nih.gov/pubmed/20675444 Rico et al., 2010]] Rico, J., Pardo, E., and Orejas, M. (2010). Enhanced production of a plant monoterpene by overexpression of the 3-hydroxy-3-methylglutaryl coenzyme a reductase catalytic domain in ''Saccharomyces cerevisiae''. ''Appl Environ Microbiol'', 76(19):6449–54.
*[[http://www.ncbi.nlm.nih.gov/pubmed/18072821 Sun, 2007]] Sun, J. (2007). D-limonene: safety and clinical applications. ''Altern Med Rev'', 12(3):259–64.
Vector Design
Thaumatin
Constitutive Promoter
Light-Switchable Promoter
Ethanol-Inducible Promoter
Genome Integration
BioBricks
Priors
Gal1 Promoter
Yeast Growth
Human Practice
Action Day
Labjournal
1 Limonene
1.1 Background and Principles
1.1.1 Biosynthesis
1.1.2 Molecular and Physiological Effects of Limonene
1.1.2.1 Flavor and Aliment
1.1.2.2 Cancer Inhibition
1.1.2.3 Solvation of Gallstones
1.2.1 BioBricks
1.2.1.1 BBa_K801060 (+)-Limonene synthase 1 with Strep-tag and yeast consensus sequence
1.2.1.2 BBa_K801061 (+)-Limonene synthase 1 coding region from Citrus limon
1.2.1.3 BBa_K801062 (+)-Limonene synthase 1 expression cassette for yeast
1.2.1.6 BBa_K801065 (+)-Limonene synthase 1 with Strep-Tag
1.2.1.7 BBa_K801066 (+)-Limonene synthase 1 with yeast consensus sequence for improved expression
1.2.2 Characterization
1.2.2.1 Gel Picture of Finished Constructs
1.2.2.2 Investigation of Yeast Consensus Sequence
1.2.2.3 Purification of Recombinant Limonene Synthase
1.2.2.4 In Vitro Detection of Limonene
1.2.2.5 In Vivo Detection of Limonene
1.2.2.6 Detection of Limonene in Beer
1.2.2.7 Toxicity Assay
Responsible: Lara Kuntz and Andrea Richter
A Common Saying Is "If Life Gives You Lemons - Make Lemonade". For Us It Is Rather "If Life Gives You Limonene - Make Beer".
Just think of the refreshing sensation of lemons paired with the complex richness of a chilled brew.
Doesn't that make you thirsty? Then you're probably an Englishman -
Just think of the refreshing sensation of lemons paired with the complex richness of a chilled tepid brew.
D-limonene is used as a component of flavorings and fragrances since it has an orange/lemon-like odor. Limonene has been shown to inhibit rat mammary and other tumor development [Tsuda et al., 2004]. Being an excellent solvent of cholesterol, d-limonene also has been used clinically to dissolve cholesterol-containing gallstones. Because of its gastric acid neutralizing effect and its support of normal peristalsis, it has also been used for relief of heartburn [Sun, 2007].
Producing the flavoring substance limonene in our beer might result in a fresh, lemon-like taste on the one hand. On the other hand, we might have beneficial effects on health such as preventive activity against cancer, dissolution of gallstones and relief of heartburn.
We have achieved our goal of a beer with limonene. We have successfully cloned (+)-limonene synthase 1 into our newly established yeast expression vector pTUM100. We transformed Saccharomyces cerevisiae with the plasmid carrying limonene synthase. We proved expression of limonene synthase via western blot. We verified the functionality of (+)-limonene synthase 1 by in vitro assay with purified limonene synthase on the one hand. On the other hand, we proved functional limonene production in yeast cell culture via headspace GC-MS. Furthermore, we analyzed differences in protein expression in yeast depending on existence of the yeast consensus sequence. To survey the toxic concentration of limonene for yeast cells, a toxicity assay was performed. We showed an inhibition of growth at 1 mM and a lethal effect at 100 mM. These concentrations should not be reached by expression of limonene synthase in yeast.
Last but not least, we have been able to proof the production of limonene in the beers we brewed. Hence, we have brewed iGEM's first SynBio beer containing limonene.
Background and Principles
Limonene is a cyclic terpene and a major constituent of several citrus oils (orange, lemon, mandarin, lime and grapefruit). It is a chiral liquid with the molecular mass of 136.24 g/mol. The (R)-enantiomer smells like oranges and is content of many fruits, while the (S)-enantionmer has a piney odor [Fietzek et al., 2001]. Therefore, d-limonene ((+)-limonene, (R)-enantiomer) is used as a component of flavorings and fragrances.
Limonene is produced by limonene synthase. Limonene synthase uses geranyl pyrophosphate (GPP), which is the universal precursor of monoterpenoids, as educt. (+)-limonene synthase from Citrus limon consists of 606 aminoacids (EC=4.2.3.20) and catalyzes the following reaction: Geranyl pyrophosphate = (+)-(4R)-limonene + diphosphate (see Fig. 1).
Fig. 1: Reaction catalyzed by limonene synthase.
Saccharomyces cerevisiae produces geranyl pyrophosphate via the mevalonate pathway (see Fig. 2). GPP occurs as an intermediate of farnesyl pyrophosphate (FPP) synthesis [Oswald et al., 2007]. It has been established that S. cerevisiae has enough free GPP to be used by exogenous monoterpene synthases to produce monoterpenes under laboratory and vinification conditions [Herrero et al., 2008, Oswald et al., 2007].
Fig. 2: Simplified isoprenoid pathway in S. cerevisiae, including the branch point to linalool. Abbreviations: HMG-CoA, 3-hydroxy-3-methylglutaryl coenzyme A; IPP, isopentenyl pyrophosphate; GPP, geranyl pyrophosphate; FPP, farnesyl pyrophosphate; DMAPP, dimethylallyl pyrophosphate; HMGR, HMG-CoA reductase; FPPS, FPP synthase; LIS, linalool synthase. [Rico et al., 2010]
Molecular and Physiological Effects of Limonene
Flavor and Aliment
Because of its pleasant citrus flavour and very low toxicity (oral LD50 for mice = 5.6 and 6.6 g/kg body weight), d-limonene is widely used as a flavor and fragrance additive. It is listed in the Code of Federal Regulations as a generally recognized as safe (GRAS) flavoring agent [FDA]. It can be found in common food items such as fruit juices, soft drinks, baked goods, ice cream, and pudding in typical concentrations of 50 ppm to 2,500 ppm [Sun, 2007]. The normal daily consume of d-limonene is 0.27 mg/kg body weight per day [Watzl, 2002]. As natural compound of plants, limonene has practical advantages with regard to availability, suitability for oral application, regulatory approval and mechanisms of action. It does not pose a mutagenic, carcinogenic or nephrotoxic risk to humans [Sun, 2007].
Cancer Inhibition
Monoterpenes show anticarcinogenic effects in animal experiments. They have been shown to inhibit rat mammary, gastric, lung and skin tumor development by several discussed mechanisms such as apoptosis induction and modulation of oncogene signal transduction [Tsuda et al., 2004, Watzl, 2002]. d-limonene induces phase I and phase II carcinogen-metabolizing enzymes such as cytochrome p450. These metabolize carcinogens into less toxic forms on the one hand. On the other hand, they prevent the interaction of chemical carcinogens with DNA. Limonene has also been shown to inhibit tumor cell proliferation, to accelerate the rate of tumor cell death and induce tumor cell differentiation. Furthermore, d-limonene regulates cell growth and/or transformation by inhibiting protein isoprenylation [Sun, 2007].
Solvation of Gallstones
d-limonene is used as excellent solvent of cholesterol, therefore it has been used clinically to dissolve cholesterol-containing gallstones [Sun, 2007]. A study with 200 patients showed that direct infusion of 20-30 ml d-limonene (97% solution) completely or partially dissolved gallstones in 141 patients. Gallstones completely dissolved in 96 cases (48%); partial dissolution was observed in 29 cases (14.5%); and in 16 cases (8%) complete dissolution was achieved with the inclusion of hexamethaphosphate (HMP), a chelating agent that can dissolve bilirubin calcium stones [Igimi et al., 1991]. Because of its gastric acid neutralizing effect and its support of normal peristalsis, it has also been used for relief of heartburn [Sun, 2007].
BBa_K801060 (+)-Limonene synthase 1 with Strep-tag and yeast consensus sequence
This part contains (+)-limonene synthase 1 of Citrus limon. It is preceeded by the yeast consensus sequence for improved expression and carries a C-Terminal Strep-Tag for purification or detection by westernblot. It is an improved version of BBa_I742111.
UniProt entry: Q8L5K3
E.C. Number: 4.2.3.20
Origin of the enzyme: Citrus limon
BBa_K801061 (+)-Limonene synthase 1 coding region from Citrus limon
BBa_K801062 (+)-Limonene synthase 1 expression cassette for yeast
This part can be used to express Citrus limon (+)-limonene synthase 1 in yeast. The expression is controlled by TEF1 promoter and CYC1 terminator.
This part can be used to express Citrus limon (+)-limonene synthase 1 in yeast. The expression is controlled by yeast TEF1 promoter and yeast TEF1 terminator.
This part can be used to express Citrus limon (+)-limonene synthase 1 in yeast. The expression is controlled by yeast TEF2 promoter and yeast CYC1 terminator.
BBa_K801065 (+)-Limonene synthase 1 with Strep-Tag
BBa_K801066 (+)-Limonene synthase 1 with yeast consensus sequence for improved expression
This part is second version of BBa_K801060. It contains (+)-limonene synthase and the consensus sequence for enhanced expression in yeast. It does not contain a C-terminal Strep-tag as BBa_K801060. The sequence does not contain a stop codon so that RFC 25 has to be used.
Gel Picture of Finished Constructs
Fig. 3: Gel electrophoresis of K801060 and K801061 after analytical restrigtion digest with EcoR1 and Pst1.
(+)-Limonene synthase 1 coding region without yeast consensus sequence [BBa_K801061] in pSB1C3 is shown next to (+)-limonene synthase 1 with Strep-tag and yeast consensus sequence [BBa_K801060] after restriction digest with EcoR1 and Pst1 restriction enzymes. To check success of ligation, DNA fragments were separated by agarose gel-electrophoresis using ethidium bromide as a nucleic acid stain. As expected, the 1665 bp fragment of [BBa_K801061] and the 1708 bp fragment of [BBa_K801060] were detected additionally to the pSB1C3 vector that has 2070 bp (see Fig. 3).
Investigation of Yeast Consensus Sequence
Fig. 4: Comparison of limonene synthase biobricks (BBa_K801065 and BBa_K801060) with and without yeast consensus sequence.
[Hamilton et al., 1987] reported of a consensus sequence upstream of the AUG start codon in yeast. Although not as strong as the mammalian Kozak translation initiation sequence, the yeast consensus sequence is thought to have a 2–3-fold effect on the efficiency of translation initiation [pYES2 manual].
We have only been able to show slight differences in expression between the two biobricks [BBa_K801065 and BBa_K801060] via coomassie staining and western blot. The difficulties of showing the difference via SDS-page may result from variations in the amount of protein applied.
Our in vivo analysis strongly indicates that the consensus sequence does lead to a 2-3-fold enhanced expression in yeast, though (see Fig. 8B). This is consistent with findings of others [pYES2 manual].
Purification of Recombinant Limonene Synthase
Fig. 5: Streptavidin Affinity (SA) chromatography.
Fig. 6: Analytical gelfiltration of limonene synthase.
Streptavidin affinity chromatography
The yeast cell extract that was obtained by cell lysis using glass beads of 0.5 mm was centrifuged at 11000 RPM in an SLA-3000 rotor for 60 minutes and subsequently dialysed against 5 liters of 1x Streptavidin Affinity buffer (SA-buffer) over night. The cell extract was then filtrated using a syringe filter with a pore size of 0.45μm and susequently applied on an SA-column. After sample application, the column was washed with 1x SA buffer until a base line was reached. Subsequently, bound protein was eluted using 5 mM Biotin in 1x SA buffer. The chromatogram of the purification is shown on the left side.
Gel Filtration of Purified Protein The protein sample obtained was concentrated using a centrifugation concentrator with a molecular size limit of 30 kDa subsequent filtration. 250μl of the solution were applied to an analytical gel filtration column Superdex 200 10/30 with 1x PBS as running buffer at a flow rate of 0.5 ml/min. In the chromatogram (shown in the figure on the right in section B), there is an aggregate peak at the exclusion limit that may be caused by the preceding concentration and a major peak at an elution volume of 13.580 ml. The calibration line that was obtained from the calibration proteins b-amylase, alcohol dehydrogenase, BSA, ovalbumine, carboanhydrase, cytochrome C and aprotinin filtrated with the same experimental setup resulted in a regression line with the formula y = -39206 x + 3.3463. Using this formula and the elution volume of the limonene synthase, an apparent molecular mass of 70.1 kDa could be determined for the produced limonene synthase. This fits quite well the theoretical molecular mass that was calculated using ExPASy ProtParam to be 65977.1 Da. The four kilo daltons difference could be caused by posttranslational modifications. This hypothesis should be tested using mass spectrometry.
In Vitro Detection of Limonene
Fig. 7: Spectrum of in vitro detection of limonene (enzyme assay with BBa_K801060) and reference spectrum.
To test the functionality of purified limonene synthase in vitro, we used an optimized protocol of an enzyme assay with extraction of limonene [Landmann et al, 2007]. The limonene synthase was purified via Strep-tag. The enzyme assay was carried out in 25 mM Tris-HCl buffer with 5% Glycerol, 1 mM DTT and cofactors (10 mM MgCl2, 1 mg/ml BSA). 50 µM substrate (geranyl pyrophosphate) and 10 ng purified recombinant limonene synthase were added to the reaction batch. Negative controls were reaction batches without enzyme. The reaction was incubated for 15 min at room temperature. Afterwards, limonene was extracted with pentane, dried with sodiumsulfate and reduced under a stream of nitrogen. Three replicates were done of both sample and negative control.
All enzyme reactions (three replicates) led to the production of limonene while the negative controls did not show limonene. Therefore, we showed that our purified limonene synthase is functional and leads to the production of limonene.
In Vivo Detection of Limonene
Fig. 8: Detection of limonene in headspace above cell culture supernatant. [A] Spectrum of limonene obtained when analyzing cell culture that was transformed with pTUM104 containing construct of BBa_K801060. [B] Overview about different measurements.
Because limonene is a VOC (volatile organic compound) [Pierucci et al., 2005], we expected limonene to be present in the gaseous phase above the cell culture as well as in the cell culture. Therefore, we measured limonene via headspace (SPME needle) GC-MS.
We showed limonene to be produced by yeast that was transformed with pTUM104 carrying limonene synthase coding regions (see BBa_K801061 and BBa_K801060).
Furthermore, we were not able to detect a significant difference between samples that had additional GPP (educt) versus the ones that did not. This might be due to the inability of GPP to diffuse into the cells (hydrophilic character). Since we were able to detect limonene in both samples, it implies that the GPP present in the cells is sufficient for limonene production. This is consistent with the findings of [Herrero et al., 2008] that showed that S. cerevisiae cells (from laboratory and wine strains) contain enough free GPP to be catalytically transformed by monoterpene synthases into monoterpenes.
Detection of Limonene in Beer
Fig. 9: Preparation of sample for GC-MS with SPME.
Fig. 10: iGEM's first and finest SynBio Beer with limonene.
A first attempt to use our genetically engineered yeasts to brew a SynBio Beer were conducted using a transient transfection with a constitutive promoter. A drawback might be that the selection pressure might not be preserved in the gyle and hence the loss of the plasmid might be possible. Therefore, we also performed brewing experiments with yeast that carried genome integrated limonene synthase.
Fig. 11: GC spectra obtained from GC-MS analysis of brewed beer. Brown: Beer brewed with yeast that carries pTUM101 with limonene synthase. Green: Beer brewed with yeast that supposedly carries limonene synthase in genome (after genome integration).
Toxicity Assay
Fig. 12: Limonene toxicity assay evaluation.
[Fietzek et al., 2001] Fietzek, C., Hermle, T., Rosenstiel, W., Schurig, V. (2001) Chiral discrimination of limonene by use of beta-cyclodextrin-coated quartz-crystal-microbalances (QCMs) and data evaluation by artificial neuronal networks. Fresenius J Anal Chem., 371(1):58-63.
[Hamilton et al., 1987] Hamilton, R., Watanabe, C. K., De Boer, A. H. (1987) Compilation and comparison of the sequence context around the AUG startcodons in Saccharomyces cerevisiae mRNAs. Nucleic Acids Res., 15(8):3581–3593.
[Herrero et al., 2008] Herrero, O., Ram ́on, D., and Orejas, M. (2008). Engineering the Saccharomyces cerevisiae isoprenoid pathway for de novo production of aromatic monoterpenes in wine. Metab Eng, 10(2):78–86.
[Igimi et al., 1991] Igimi, H., Tamura, R., Toraishi, K., Yamamoto, F., Kataoka, A., Ikejiri, Y., Hisatsugu, T., Shimura, H. (1991). Medical dissolution of gallstones. Clinical experience of d-limonene as a simple, safe, and effective solvent. Dig Dis Sci., 36(2):200-8.
[Landmann et al., 2007] Landmann, C., Fink, B., Festner, M., Dregus, M., Engel, K.-H., and Schwab, W. (2007). Cloning and functional characterization of three terpene synthases from lavender (Lavandula angustifolia). Arch Biochem Biophys, 465(2):417–29.
[Lücker et al., 2002] Lücker, J., El Tamer, M. K., Schwab, W., Verstappen, F. W. A., van der Plas, L. H. W., Bouwmeester, H. J., and Verhoeven, H. A. (2002). Monoterpene biosynthesis in lemon (Citrus limon). cDNA isolation and functional analysis of four monoterpene synthases. Eur J Biochem, 269(13):3160–71.
[Oswald et al., 2007] Oswald, M., Fischer, M., Dirninger, N., and Karst, F. (2007). Monoterpenoid biosynthesis in Saccharomyces cerevisiae. FEMS Yeast Res, 7(3):413–21.
[Pierucci et al., 2005] Pierucc, P., Porazzi, E., Martinez, MP., Adani, F., Carati, C., Rubino, FM., Colombi, A., Calcaterra, E., Benfenati, E. (2005). Volatile organic compounds produced during the aerobic biological processing of municipal solid waste in a pilot plant. Chemosphere, 59(3):423-30.
[Rico et al., 2010] Rico, J., Pardo, E., and Orejas, M. (2010). Enhanced production of a plant monoterpene by overexpression of the 3-hydroxy-3-methylglutaryl coenzyme a reductase catalytic domain in Saccharomyces cerevisiae. Appl Environ Microbiol, 76(19):6449–54.
[Sun, 2007] Sun, J. (2007). D-limonene: safety and clinical applications. Altern Med Rev, 12(3):259–64.
[Tsuda et al., 2004] Tsuda, H., Ohshima, Y., Nomoto, H., Fujita, K., Matsuda, E., Iigo, M., Takasuka, N., Moore, MA. (2004). Cancer prevention by natural compounds. Drug Metab Pharmacokinet. 19(4):245-63.
[Watzl, 2002] Watzl, B. (2002). Monoterpene. Ernährungs-Umschau 49 Heft 8. 322-324.
[Williams et al., 1998] Williams, D. C., McGarvey, D. J., Katahira, E. J., and Croteau, R. (19
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Vol. 11, Issue 25 - Wednesday, January 25, 2006
W | T | F | S | S | M | T
» Vital Statistics
Joan Louise Barnette
Joan Louise Barnette, 72, of Mountain View, Hawaii, a bartender, died in Hilo Medical Center. She was born in Oregon. She is survived by husband Dan, sons James and Russel Brand and Brian Bittner, six other children, 22 grandchildren, and five great-grandchildren. Services at a later date.
Roy Clayton Boehm
Roy Clayton Boehm, 81, of Honolulu, a retired federal Immigration and Naturalization Service assistant district director for investigation in Los Angeles, died in Straub Clinic & Hospital. He was born in Orofine, Idaho. He is survived by wife Robyn; sons Roy L. Boehm and Jonathan K. White, daughters Julie Hess and Beurlie U. Christensen, eight grandchildren and five great-grandchildren. Services: 11 a.m. Saturday at Scottish Rite Cathedral, 1611 Kewalo St. Call after 10 a.m. Private inurnment at National Memorial Cemetery of the Pacific, Punchbowl. Casual attire. No flowers.
Sau Yee Chong
Sau Yee Chong, 90, of Hono-lulu, died. She was born in Honolulu. She is survived by daughter Yvonne Lee, brother Wah Quon Leong, three grandchildren and six great-grandchildren. Private services.
Frank Joseph Correnty
Frank Joseph Correnty, 85, of Kihei, a retired Exxon Oil Co. vice president and Army veteran who served in World War II, died at home. He was born in New Jersey. He is survived by companion Beverly Smith, daughter Diane Tower and two grandchildren. Services: 11 a.m. Friday at Ballard Family Mortuary. Call after 9 a.m. Burial: noon at Maui Veterans Cemetery.
Janella Lei Tabua Daguio
Janella Lei Tabua Daguio, infant daughter of Edimar and Lenie of Waipahu, died in Kapiolani Medical Center for Women & Children. She was born in Honolulu. She is also survived by sister Jessanelle and grandparents Gaudencio Evaresto Sr. and Angelina Daguio, and Bienvenido and Armelina Tabua. Services: 10:30 a.m. Saturday at Mililani Mortuary-Waipio, makai chapel. Call after 8:30 a.m. Burial: 11 a.m. at Mililani Memorial Park. Casual attire.
Ismael Epil
Ismael Epil, also known as "Blackie" and "John," 72, of Waimanalo, a retired Army National Guard member and Castle Medical Center maintenance technician, died in Honolulu. He was born in Waimanalo. He is survived by mother, Saturnina, son Michael, daughter Joslyn Imura, brother Juanito, sister Sister Charlene and three grandchildren. Services: 11:45 a.m. Friday at St. George Catholic Church. Call after 9:30 a.m. Graveside services: 10 a.m. Tuesday at Hawaii State Veterans Cemetery, Kaneohe. Aloha attire. No flowers.
Beverly Y. Feleti
Beverly Y. "Angel" Feleti, 50, of Mililani, a salesclerk, died in Wahiawa General Hospital. She was born in Honolulu. She is survived by husband Tufulemataafa, sons Cyrus Edwards and Tufulemataafa M. Feleti, daughters Michelle White and Ianeta B. Feleti, brothers Able and Bernard Lui, sisters Brenda Garrigan and April Phua, and two grandchildren. Services: 1 p.m. Sunday at Oahu Cemetery Chapel, 2162 Nuuanu Ave. Call after noon. Burial: 11 a.m. Monday at Hawaii State Veterans Cemetery, Kaneohe. Casual attire.
Ronald O'Sullivan Fernandez
Ronald O'Sullivan Fernandez, 72, of Pearl City, an Associated Steel Workers Co. iron worker, died at home. He was born in Ualapue, Molokai. He is survived by wife Cecilia, son Ronald Jr., daughters Clarine Hipa and Cara-Lynne Fernandez, brother Lot Charles, sister Lehua and five grandchildren. Services: 10:30 a.m. Friday at Borthwick Mortuary. Call after 9 a.m. Burial: 2 p.m. at Hawaii State Veterans Cemetery, Kaneohe. Casual attire.
Hugo Mark Garcia
Hugo Mark Garcia, 50, of Honolulu, a computer programmer, died in Honolulu. He was born in Lima, Peru. He is survived by mother Yolanda, brothers Omar and Luis, and sisters Miriam Vermejo and Toti Garcia. Services: 5 p.m. Sunday at Kilohana United Methodist Church. Casual attire. No flowers.
Clara Chieno Enomoto Ito
Clara Chieno Enomoto Ito, 58, of Honolulu, a retired teacher, died in Honolulu. She was born in Honolulu. She is survived by husband Clifford R. and daughter Erin. Services: 6 p.m. Saturday at Nuuanu Memorial Park & Mortuary, east chapel. Call after 4:30 p.m. Private inurnment at a later date.
James Ho'omaika'i Kaauwai
James Ho'omaika'i Kaauwai, 67, of Honolulu, owner of Jimmy's Appliance, died in Tripler Army Medical Center. He was born in Lihue. He is survived by daughters Kuuipo Kaauwai-George, Pua Kaauwai and Jessica Yaple; brothers John, William and Bert; sisters Elizabeth Costa, Henrietta Gonsalves, Maureen Seagrove, Annette Kaauwai and Linda Iwamoto; and five grandchildren. Services: 11 a.m. Saturday at Kapaa First Hawaiian Church. Call after 9 a.m. Cremation to follow. Casual attire. Leis only.
Terry K. Kamalamalama Sr.
Terry K. Kamalamalama Sr., 51, of Makakilo, a professional musician and singer, died in Straub Clinic & Hospital. He was born in Hilo. He is survived by wife Noenoe; sons Abram R. "Keao," Terry "Kalei" Jr. and Noah; brothers George, Joseph and Bernard; and sisters Alberta and Wilhemina. Services: 4 p.m. Saturday at the Church of Jesus Christ of Latter-day Saints, Makakilo Stake, 92-900 Makakilo Drive. Call after 1 p.m. Aloha attire.
Takayo Kobayashi
Takayo Kobayashi, 100, of Honolulu, a retired Japanese language school teacher, died in St. Francis-Liliha Medical Center. She was born in Hono-mu, Hawaii. She is survived by husband Torao; daughter Marjorie F.; sons Bert N., Lawrence A., Arthur T. and Gary S.; sisters Jane Nakayama, Yukie Hayashi and Sachiko Toyofuku; nine grandchildren; and eight great-grandchildren. Services: noon Sunday at Hosoi Garden Mortuary. Casual attire. No flowers.
Sarah Frances Gertz Marion
Sarah Frances Gertz Marion, 87, of Lanikai, an artist and teacher, died in Lanikai. She was born in Honolulu. She is survived by son Gregg, daughters Ann Marion and Donna Rasmussen, 12 grandchildren and seven great-grandchildren. Graveside services: 10:30 a.m. Saturday at Diamond Head Memorial Park.
Ben Bruce Mayes
Ben Bruce Mayes, 78, of Waianae, died in Kaiser Medical Center. He was born in Huntsville, Texas. He is survived by wife Florence M., son Chad H. Omori, daughter Lisa K. Jones and two grandchildren. Services: 4 p.m. Saturday at Mililani Mortuary-Waipio, makai chapel. Call after 3:30 p.m. Casual attire.
Joseph Albert Miller III
Joseph Albert "Milla" Miller III, 58, of Honolulu, died at home. He was born in Hono-lulu. He is survived by sisters Georgia Vager, Venus Cabinatan and Vallery Lau; brothers Edward and Rudolph Sun; hanai brother Ernest Wright; and hanai daughter Marvalee Wright Lin Kee. Visitation: 6 to 9 p.m. Monday at Hawaiian Memorial Park, 45-425 Kamehameha Highway, Kaneohe. Scattering of ashes at a later date. Leis only.
Betty Sumiko Miyamoto
Betty Sumiko Miyamoto, 84, of Honolulu, a retired Kahala Sportswear seamstress, died in Hale Ho Aloha. She was born in Honolulu. She is survived by sons Calvin K., Roy S. and Eric K.; daughter Susan C. Oda, sister Dorothy Shimomaye and six grandchildren. Services: 4 p.m. Sunday at Hosoi Garden Mortuary. Call after 3 p.m. Casual attire. No flowers.
Roy Seichi Nakamoto
Roy Seichi Nakamoto, 92, of Honolulu, retired owner of Roy S. Nakamoto Plumbing, died in Hale Nani Rehabilitation and Nursing Facility. He was born in Haiku, Maui. He is survived by wife Misae, daughters Kay S. Uyeda and Shirley M. Nakamoto, and brothers Kenji and Tadashi. Services: 4 p.m. Monday at Hosoi Garden Mortuary. Casual attire. No flowers.
Neil Hiro Nakao
Neil Hiro Nakao, 47, of Pearl City, died in Aloha Nursing & Rehabilitation Centre. He was born in Honolulu. He is survived by parents Yasuo and Doreen, and sisters Lori Ann Nakao-Yamada and Jani Nakao-Otaka. Services: 6:30 p.m. Friday at Mililani Mortuary-Waipio, makai chapel. Call after 6 p.m. Casual attire. No flowers. Incorrect information was published in an obituary yesterday.
Michael S. Neill
Michael S. Neill, 52, of Sonora, Calif., died in Twain-Harte, Calif. He was born in California. He is survived by father Don, mother Janice Shea and sister Jamie. Services: 9 a.m. Sunday at Kailua Middle Beach Park (across Buzz's Steak House). No flowers. Donations suggested to charity.
Edgardo Pangilinan
Edgardo Pangilinan, 64, of Koloa, Kauai, a retired Sheraton groundskeeper, died in Wilcox Memorial Hospital. He was born in Manila. He is survived by wife Raquel; son Randy; brother Nicanor; and sisters Adelina Maggay, Rosario Lacuesta, Elisa Nerida and Susuana Casi. Services: 11 a.m. Friday at Borthwick Kauai Mortuary Chapel, Koloa. Call after 9 a.m. Burial to follow at Koloa Public Cemetery. Casual attire.
Edward N. Rabot
Edward N. Rabot, 73, of Kapaa, Kauai, a retired Shimamoto Contractor carpenter, died at home. He was born in Koloa, Kauai. He is survived by sons Edward A., Edward and Jason; daughters Celestine Negley, Dawn A. Rabot and Charmaine Penney; brothers Henry N. and Francisco Jr.; sisters Cecelia Gabriel, Linda Enos and Rosalina Rabot; and eight grandchildren. Services: 11 a.m. Friday at Garden Island Mortuary Chapel. Call after 9 a.m. Burial to follow at Kauai Veterans Cemetery. Casual attire.
Georgia-Ann Lani Spencer Remmers
Georgia-Ann Lani Spencer Remmers, 51, of Pahoa, Hawaii, died in Hilo Medical Center. She was born in Hilo. She is survived by daughters Leigh-ann A.B.K. and Dayna-lyn I.K. Spencer, Rynae M. Mendes and Angela L. Kaawaloa; brothers Walter, Rudy and Bryant Spencer; sisters Lavaina Lindsey and Hawley Reese; and 12 grandchildren. A potluck memorial service will be held noon Sunday at Carvalho Park, Hilo.
Audrey Rowe
Audrey "Molly" Rowe, 83, of Lihue, an accountant, died in Wilcox Memorial Hospital. She was born in Wisconsin. She is survived by husband Bernard W., sons Dennis W. and Robert S., daughter Christine E. Taniguchi, 10 grandchildren and 15 great-grandchildren. Services: 11 a.m. tomorrow at Lihue Lutheran Church. Call after 10 a.m. Inurnment to follow at the church cemetery. Casual attire.
Betty Sadae Tanaka Santiago
Betty Sadae Tanaka Santiago, 86, of Wahiawa, a retired Tripler Army Medical Center employee, died. She was born in Aiea. She is survived by sons Charles, Victor "Sandy," Kenneth and Randall "Randy"; daughter Susan "Sue" Sheldon; brother Thomas "Tommy" Tanaka; sister Florence "Aunty Koga" Koga; 18 grandchildren; and 15 great-grandchildren. Services: 6:45 p.m. Sunday at Borthwick Mortuary. Call after 5 p.m. Mass: noon Monday at Cathedral of Our Lady of Peace. Call after 10 a.m. Burial: 1 p.m. at National Memorial Cemetery of the Pacific, Punchbowl. Aloha attire.
Alice Louise Judd Simpich
Alice Louise Judd "Awee" Simpich, 87, of Kula, Maui, a sculptress and homemaker, died at home. She was born in Honolulu. She is survived by daughter Louise B. "Louli" MacCluer and four grandchildren. Graveside services: 4 p.m. Friday at Makawao Cemetery. No flowers. Donations suggested to Maui Humane Society or charity.
Happy Smith
Happy Smith, 32, of Kahului, a Bradley Pacific fueler, died in Maui Memorial Medical Center. He was born in Micronesia. He is survived by fiancée Delianna Defang, son Donavon Defang, daughter Lokilani and sister Michaela. Services: 5 p.m. Saturday at Iao Congregational Church. Call after 4 p.m. Burial in Micronesia.
Ramon Soriano
Ramon Soriano, 46, of Ewa Beach, died in the Philippines. He was born in San Carlos City, Pangasinan, the Philippines. He is survived by wife Roselle; sons Ishbel, Seth and Zachary; brother Joselito; and sisters Gina Tagawa and Angie Pineda. Services: 10:30 a.m. Monday at the Church of Jesus Christ of Latter-day Saints, Ewa Beach 2nd Ward. Call after 8:30 a.m. Burial: 2 p.m. at Hawaii State Veterans Cemetery, Kaneohe.
Tomiko Kapualani Tamura
Tomiko Kapualani Tamura, 64, of Ewa Beach, died in Kapiolani Medical Center, Pali Momi. She was born in Kapaa, Kauai. She is survived by husband Reginald S. "Moose," son Blaine J.K.S., daughters Diane T. Gussman and Annette S. Clark, brothers Jacob Martin Sr. and Kalani Tachibana, sisters Minerva DePonte and Dorothy Smith, four grandchildren, and two great-grandchildren. Services: 11:30 a.m. Saturday at Mililani Mortuary-Waipio, mauka chapel. Call after 9 a.m. Burial: 12:30 p.m. at Mililani Memorial Park. Aloha attire.
Satoshi Tanita
Satoshi "Jimmy" Tanita, 82, of Honolulu, a retired self-employed plumber doing business as Central Plumbing, died in Kaiser Medical Center, Moanalua. He was born in Waimea, Kauai. He is survived by wife Lillian T.; sons Roddy and Russell; daughters Joyce Nakatani, Lorrie Iwanaga and Gail Pine; brothers Tamotsu and Mitsuyoshi Hiraoka; sister Koharu Kawamura; and seven grandchildren. Services: 2 p.m. Sunday at Hosoi Garden Mortuary. Call after 1 p.m. Aloha attire.
Damian Kekoa Draco Weldy
Damian Kekoa Draco Weldy, infant son of Jonathan Weldy and Tianna Moore of Waianae, died in Waianae Coast Comprehensive Health Center. He was born in Honolulu. He is also survived by grandparents Johnny and Pamela Weldy, and David and Lynn Moore; and great-grandparents T. & June Moore and Austin Clough. A memorial gathering will be held 10 a.m. Friday at Makua Beach Park.
You can also search the Hawaii State Library System's Hawaii Newspaper Index online for older obituaries at www.librarieshawaii.org. The index goes back several decades.
» Gas cap repeal gains steam
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» Delayed environmental review at heart of Makua conflict
Betty Shimabukuro
Paul Arnett
Cynthia Oi
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Academic Recognition Banquet features speakers Allison and Amanda Lansell
The 18 students that make up the top ten percent of the graduating class of 2018 were honored, along with their honored educator of choice and their parents at the annual Academic Recognition Banquet on Wednesday evening.
The purpose of the Academic Recognition Day is to publicly honor the academic top ten-percent of Alliance’s graduating senior class, their parents/guardians, and the educators whom the students feel have most influenced their lives. The dinners have been an annual tradition for 31 years.
This year’s featured speakers were Allison and Amanda Lansell. The twins graduated in the Alliance High School class of 2005 and are now a lawyer and doctor, respectively.
Allison attended the University of Virginia Law School and graduated in 2012. Upon graduation she moved to Washington DC to begin her career. She is licensed to practice law in Virginia, Maryland, Washington DC, and Ohio. Her law practice has varied from civil to criminal work, including both prosecution and defense. She now advises nursing homes in a wide array of legal needs. She lives and works in the Cleveland area and has been named to Super Lawyers Rising Stars for Health Law the past two years.
Amanda graduated in 2013 from the University of Toledo College of Medicine. She completed a 3-year Pediatrics residency at Rainbow Babies and Children’s Hospital in Cleveland, Ohio and a 1-year Hospital Medicine fellowship at Emory University in Atlanta, Georgia, winning awards for her teaching at both institutions. She currently works as a pediatric hospitalist, caring for hospitalized children and delivering newborn babies at three different Rainbow Babies and Children’s Hospitals in the Cleveland area.
The two spoke during the dinner at the University of Mount Union and encouraged students to press on through failure in order to find success.
“It is hard to talk about your failures. I want you to know its ok to fail. You are going to fail at some point. When you are faced with failure, let it deter you from goals or use it to motivate you even more,” Amanda said.
Both told about failures they have had in their own careers and what they had to do to keep going in order to reach success.
“How you respond to failure defines you…Sometimes we fail despite our best efforts. How you move forward is what matters,” Allison said.
Each student at the dinner received a $200 check from the Alliance Foundation for their education. Howard Frazier also received a $500 scholarship from the John and Roene Klusch Foundation for earning the top ACT score in the group.
The event is sponsored each year by the John and Roene Klusch Foundation.
John and Roene Klusch were life-long residents of Alliance, Ohio and members of the St. Paul Lutheran Church in Alliance. John was the founder of East Ohio Machinery Company, which still operates in Alliance Ohio. The couple established the Foundation to support Alliance, Ohio and Marshallville, Ohio, where Roene was raised. In addition to the Foundation, the Klusch’s were strong supporters of the Alliance High School Scholarship Fund.
The following students and educators were recognized at the banquet:
Estella Bennett
To attend The University of Mount Union and study psychology.
Kiara is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Member of National Honor Society for 2 years
Member of Stark County Teen Court for 3 years
Member of AHS Girls Soccer Team for 3 years
Member of AHS PenOhio Writing Team for 3 years
Recipient of Aultcare Team Spirit Award
Volunteer at Green Village Nursing Rehab Center for 2 years
Received the US Scholastic Excellence Award
Received the Stark County Helping Hand Award
Received the Stark County Teen Court Outstanding Overall Award
HONORED EDUCATOR:
Ms. Jennifer Avers, Art Instructor for 4 years
In art class, Ms. Avers taught me to overcome obstacles with these words that I have used for my life as well. “ In your projects you will reach a point that you are 75% of the way there. Things are going good and you will hit a wall where you will want to stop and give up. Don’t give up. If you push through, your art will be a success. You just have to keep going.” I will carry this with me forever.
Stephanie Alexi and Terry Bert
Attend the University of Mount Union and major in marketing.
Natalie has been awarded an Investment Alliance Scholarship from the University of Mount Union.
Member of National Honor Society for 1 year.
Member of AHS Girls Golf Team for 3 years.
Church Soloist at Alliance First Church of God for 8 years.
Youth Assistant for 8 years at Alliance First Church of God.
As a senior at AHS has attended the University of Mount Union full time as a college credit plus student.
Corrina Brindak, Youth Director at Alliance First Church of God
Corrina has known me since I was a child and has taught me so much out of the classroom.
Mary and Marty Cvelbar
To attend The University of Mount Union and study English and political science.
Mara is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Member of National Honor Society for 2 years.
Member of AHS Girls Varsity Soccer Team for 4 years.
Member of AHS Girls Swimming Team for 4 years.
Member of AHS Girls Softball Team for 4 years.
Member of Student Senate for 4 years; Student Senate President during senior year.
Member of Key Club for 4 years
Member of Foreign Language Club for 4 years
Vice President of the Class of 2018
Member of Marching Band and Flagline
Mrs. Deirdre Coar, Soccer Coach
I admire Mrs. Coar so much as a person, teacher and coach. She knew the needs of every individual and helped every single person while not playing favorites. She has taught me to not only work hard, but to be kind while doing so.
Lori Fountain and Ambrose Smith
To attend Kent State University in Salem to study Early Childhood Education.
Member of AHS Social Justice Team for 2 years
Member of University of Mount Union Together Initiating Excellence Program for 4 years
Member of AHS Foreign Language Club for 4 years
Member of Marching Band, Pep Band and Concert Band for 4 years
Member of AHS Peer Pilot Program for 2 years
Member of Teen Court for 2 years
Elks Teen of the Month for November, 2017
2018 President of the Band, Section Leader, Squad Leader
1st place winner of History Department Essay contest “There Ought To Be A Law”
Mrs. Renee Barnes, French Teacher
Mrs. Barnes has significantly impacted my life. She has always been kind, generous and always willing to help. I know I can go to her about anything and I am grateful for the opportunity to have her as a teacher.
Kim and John Frazier
To attend John Carroll University to major in Political Science and play football and then continuing my education in law school.
Member of Academic Challenge Team for 4 years
Member of AHS Varsity Football Team for 4 years
Member of AHS Varsity Track Team for 4 years
Sports Editor for Red & Blue Flyer for 3 years
Member of Choralaires for 3 years
Recipient of All-Star Preps Excellence in Action Award
Recipient of National Football Foundation Scholar Athlete Award
Published writer in “It’s My Alliance” Magazine
Member of Regina Coeli Catholic Church
Mr. Wilson works to foster meaningful discussion. However, he never becomes polarized himself and is accepting of everyone.
Lori and John (Buddy) Gasparik
To attend the University of Mount Union and major in Nursing.
Madison is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Member of Student Senate for 4 years
Scholar Athlete for 4 years
Elks Teen of the Month for January, 2018
Alliance Review Athlete of the Week
Received 1st Team All League in Golf during junior year
Received 1st Team NBC in Golf for junior and senior years
Member of Stark County Youth Ministry Board for 3 years
Recipient of Eagle of the Cross Award senior year
Ali Myslinski, 7
Miss Myslinski has been an inspiration to me. She believed in me, when I didn’t believe in myself, and she pushed me to be the best I could be.
Mary and Shawn Jackson
To attend Ohio University and major in Journalism.
Member of AHS Softball Team for 4 years
Member of AHS Volleyball Team for 4 years
Member of AHS Jetsetters for
Member of AHS Marching Band
Secretary for the Class of 2018
Participant at Girls State
Participant at Hugh O’Brien Youth Leadership Conference in 10
Amy Miller, Anatomy & Physiology, Medical Terminology and AP Biology Teacher
Mrs. Miller has always been the teacher that pushes me to be the best I can be. She has made me the hard-working student I am today.
Alix Kelley
Debbie and Ted Kelley
To attend the University of Mount Union with an undecided major
Alix is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
4 year varsity letter winner for Girls Tennis
1st Team all NBC for Tennis senior year
2 year varsity letter for Softball
Member of Marching, Concert, Pep and Jazz Bands for 4 years.
Member of OMEA District 8 Honor Band during senior year
Volunteer at Trinity Gospel Temple
DeAnn Zavarelli, 6
Mrs. Zavarelli taught me to always stand up for my beliefs. She truly cares for and respects every one of her students, and that is something I will never forget.
Danielle and Christopher Marazza
To attend The Ohio State University and major in Accounting
Kelly Trout, Biology and Chemistry teacher
Mrs. Trout made science classes that I did not have a passion for enjoyable. She is also very easy to approach and I was never reluctant to ask for help outside of class time.
Vicki and Keith McGeehen
To attend Duke University and major in Economics or Political Science
Brendan is the recipient of the Elvin J. Wilson Scholarship to Duke University
Member of Red & Blue Flyer for 4 years, Co-Editor for 3 years
Member of Football Team for 4 years
Participant at Hugh O’Brien Youth Leadership Conference in 10Active volunteer at Alliance Food Pantry
Attended Rotary Youth Leadership Conference
Received Best in Category for Engineering at Science Alliance during junior year at AHS
Richard Morris, Grandfather and Pre-Calc Teacher
His genuine love for math rubbed off on me and pushes me to become the best math student I can be.
Teresa and Steve McMillen
To attend The University of Mount Union and NEOMED to study Pharmacy
Andrew is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Member of Student Senate for 1 year
Employed at Hartville Hardware
Active Volunteer at the Stark County Humane Society and the Salvation Army
Member of AHS Orchestra for 4 years
Received excellent and superior ratings at the OMEA State Orchestra event for 4 years
Member of Varsity Soccer Team for 3 years
Member of Varsity Tennis Team for 4 years
HONORED EDUCATOR: Thomas Wilson, High School English Teacher
Mr. Wilson truly cares about his students and their success. He has been a great role model in my life as well. Thank you for all that you do.
Kristi and Mark Moore
To attend Walsh University and major in Criminal Justice
Member of Band for 4 years
Member of Orchestra for 4 years
Member of Girls Golf Team for 3 years
First Team All Stark County Conference for Bowing for 2 years
Member of Bowling Team for 4 years
Top Female Bowler for 4 years
HONORED EDUCATOR: David Reith, Teacher in 3rd, 4
I had the opportunity to have Mr. Reith in 3rd, 4 He always encouraged me to do my best and he always made learning fun.
PARENTSS:
Teri Kirby and Joseph Novak
To attend Rochester Institute of Technology and major in photography.
President of Class of 2018 for 4 years
Seamstress at Henri’s Cloud Nine
Jamie Randolph, Middle School math teacher
Mrs. Randolph was always honest and open when it came to school and life in general.
Lexi Orzo
Erin Briggs and Brian Orzo
To attend The University of Mount Union with an undecided major.
Lexi is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Recipient of Career & Tech Home Award
Member of RAM Ministries for 3 years
Member of First Friends volleyball
Member of Regina Coeli youth group
HONORED EDUCATOR: Allison Morrison, High School Guidance Counselor
Mrs. Morrison has had a lasting impact on my education because she always encouraged me to try harder and to push myself to achieve more.
Ron Rogers
To attend The University of Mount Union and major in computer science.
Cortaise is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Member of 992016 MLK Parade King
Member of Varsity Bowling Team for 3 years
State Qualifier in Bowling in 11Ira G. Turpin Scholar in 112017/18 East-West All Star Bowling Match champion
Received Honorable Mention in bowling for 2 years
Employed as Crew Trainer at McDonalds for 2 years
Participated in National Day of Reading at Northside Intermediate for 2 years
HONORED EDUCATOR: Aaron Hill, Varsity Bowling Coach
Mr. Hill has always been more than a coach or educator. He has motivated me to be my best, even when I’m at my worst. He is truly an amazing person.
Laura and Donnell Sams
Maria is a recipient of a Founders Scholarship and a 2018 Academic Challenge Scholarship from the University of Mount Union.
President of National Honor Society senior year
Member of Girls Tennis Team for 4 years
Student Representative to the Alliance City Schools Board of Education
Delegate for Buckeye Girls State in 2017
Member of Teton Science Club
4 year Scholar Athlete
Employed at Hot Pots for 3 years
6 year active volunteer and service through RAM Ministries and Regina Coeli Youth Group
HONORED EDUCATOR: Dan Dennis, High School Health Teacher
Mr. Dennis made a lasting impact on my life through his teaching due to his excellent ability to relate real life to the subject matter. There was never a day I left his class feeling that I hadn’t learned something valuable.
Lin Wu and Liang Shi
To attend the University of Cincinnati, Conservatory of Music, with a major in ballet and minor in entrepreneurship and mathematics. She will be a member of the Darwin T. Turner Scholars Program
Erika is a recipient of a Cincinnatus Century Scholarship, the Darwin T. Turner Scholarship and Buxbaum Workum Scholarship from the University of Cincinnati
Delegate at 2017 Buckeye Girls State
Received the achievement Award for Excellence in Pre-Calculus
Received the outstanding mathematics Award for Excellence in Algebra 2 Honors.
Received Canton Ballet’s Barre Association Summer Study Scholarship
Received scholarships to the Philadelphia Dance Theatre Summer Program, Carolina Ballet Summer intensive and Joffrey New York summer intensive.
Performed as Sugar Plum Fairy, Snow Queen, Dew Drop and Lead Marzipan in the Canton Ballet’s “The Nutcracker”
Performed with the Canton Ballet for 10 years
HONORED EDUCATOR: Laura Bryan, High School Guidance Counselor
Mrs. Bryan has always been understanding of my passion and dedication to ballet. Her constant support and energy have helped me through my four years of high school and two sentences are not enough to show my gratitude and respect for her. Thank you Mrs. Bryan for shaping me into the person I am today.
NAME: Maddie Waggoner
PARENTS: Amy and Edward Waggoner
PLANS: To attend the University of Mount Union and major in Exercise Science and then obtain a Doctorate of Physical Therapy
AWARDS: Maddie is a recipient of an Investment Alliance Scholarship from the University of Mount Union.
Class Treasurer for the Class of 2018 for 3 years
Member of Marching Band for 4 years
Nominee for the 2018 US Presidential Scholar in Career & Tech
All Star Preps Best of Stark County for Softball in 2017
NET Tournament Second Doubles Champion in Tennis as an 11Member of AHS Varsity Softball Team for 4 years
Member of AHS Varsity Tennis Team for 3 years
HONORED EDUCATOR: Shaun Fontaine, Band Teacher for 7 years
Mr. Fontaine has taught me to not be afraid of failure and always pushed me to be my best, as a musician and a person. He believed in me when I didn’t believe in myself.
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Showing posts from March 20, 2018
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Volkswagen Atlas Five-Seater Concept
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Mercedes-AMG GT R new pace car in the 2018 Formula 1 season
The most powerful Official FIA F1 Safety Car of all timeAffalterbach. Mercedes-AMG will use the most powerful Official FIA F1 Safety Car of all time in the 2018 Formula 1® season: the 430 kW (585 hp) AMG GT R will have its debut as the Official FIA F1 Safety Car of the FIA Formula 1 World Championship at the Grand Prix in Melbourne, Australia from 25 to 27 March 2018. The top-of-the-line model of the AMG GT series will ensure maximum safety for the Formula 1 field on all 20 race weekends. The performance and sports car brand from Affalterbach has supplied the pace car in the premier racing series of international motorsports since 1996. With an output of 430 kW (585 hp) and a top speed of 318 km/h, the AMG GT R is the most powerful and fastest Official FIA F1 Safety Car of all time: In this, the 23rd consecutive year, Mercedes-AMG will safely lead the Formula 1 field around the track when bad weather or accidents require its deployment. The AMG GT R is the ideal Official FIA F1 Safet…
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Lukashenka Compromises To Gain Moscow’s Support
On December 19, ten candidates will contest the Belarusian presidency. The election has been notably open, with several opposition candidates reaching out for support from Moscow, and the European Union offering encouragement to the incumbent president conditional on some basic requirements for a democratic process. However, belatedly the president has reached a pre-election agreement with Moscow on oil and gas imports to gain qualified support, at a high price. Opinion polls suggest that although Alyaksandr Lukashenka leads his rivals, he lacks a clear majority
Lukashenka’s electoral rating two weeks before the election was between 31 and 48 percent. Indeed, Lukashenka’s closest rivals trailed: Uladzimir Nyaklayeu, leader of the “Speak the Truth” campaign on 15.1 percent to 16.8 percent, Andrei Sannikau, 8.6 percent to 10.6 percent, and Yaraslav Ramanchuk a respectable fourth (6.1 percent to 8.2 percent), with the remainder of the field well behind. According to the poll conducted by the Ukrainian agency SOCIUM, approximately one-third of voters were still undecided (http://nekliaev.org/en/news/actual/socopros-v-belorussii-lukashenko-vinoven-453-ego-prezidentskiie-reieting-314.html, www.belmy.by, December 8).
The recent rift between Belarus and Russia appeared to make Lukashenka more vulnerable. Moscow made it clear that it would not necessarily guarantee another “elegant” Lukashenka victory and last September on his video-blog, President Dmitry Medvedev commented that the Belarusian leader had broken all the rules of decency (www.rt.com, October 4). The language was ominously similar to the letter with which he severed relations last fall with former Ukrainian President, Viktor Yushchenko (Interfax, August 11).
The Belarusian authorities had to act promptly to censor several Russian television programs that attacked and ridiculed their president. However, Lukashenka received unexpected backing from the Lithuanian President, Dalia Grybauskaite (www.charter97.org, November 23), and after a November visit to Minsk, Guido Westerwelle and Radoslaw Sikorski, the Foreign Ministers of Germany and Poland respectively, informed Lukashenka that if the election was conducted fairly, they would use their influence to solicit a $3.8 billion EU loan for Belarus, with the assistance of the International Monetary Fund (IMF) (www.thenews.pl, November 22).
Thus, the election campaign for several weeks turned Belarusian politics on its head. The EU appeared to be behind Lukashenka, while the opposition candidates, particularly Sannikau and Neklyayeu appealed to Russia. However, Moscow declined to endorse any of the nine candidates, despite support for them at lower levels of the Russian hierarchy. It left the door open for negotiations in the knowledge that Lukashenka would be in Moscow in early December for talks on the introduction of a common economic space (along with Kazakhstan), as well as meetings of the Eurasian Economic Community, the Collective Security Treaty Organization (CSTO) and the CIS (Belorusskiy Partizan, December 10).
While paying some lip service to a democratic election, the president distanced himself from “the circus.” Lukashenka informed a German audience that he expected a “landslide” victory and that the entire opposition between them could only amass about 1.5 percent (RTT News, November 15). The dismissive tone echoed his mid-September comment (a familiar one) that the Belarusian oppositionists were “enemies of the people” (RIA Novosti, September 16). Meanwhile, Lukashenka increased the average wage to $500 per month and promised it would increase to $1,000 by 2015 (SB Belarus’ Segodnya, December 7).
During the December “debate” on Belarusian Television, Lukashenka declined to appear. The program announced that the nine candidates offered identical programs –a comment repeated by Lukashenka a few days later (SB-Belarus’ Segodnya, December 8). Nyaklayeu walked out in disgust (Nasha Niva, December 4). Yet, the opposition candidates did not help their own cause. Sannikau and Statkevich appealed to viewers to gather on the “square” (Kastrichnitskaya Square in the center of Minsk) at 8.00 pm on December 19, without giving specific reasons for doing this. Despite some friendly words and cooperation between the Nyaklayeu and Sannikau camps, the opposition failed to unite behind a common candidate: personal ambitions took priority.
Lukashenka’s All-Belarusian Popular Assembly was held directly after the TV debate on December 6-7, with 2,500 selected delegates from different sectors of society, who received gifts including a watch and microwave oven, as well as having all their travel and accommodation paid at the state’s expense (Nezavisimaya Gazeta, December 6). In return, they had to endure several Lukashenka speeches based on his election platform, published in the newspaper Respublika (November 26), about the success of the economy and his future plans. Lukashenka then flew to Moscow.
Unexpectedly, a private meeting between Medvedev and Lukashenka took place immediately after the Interstate Council of the Eurasian Economic Community, during which both sides made conciliatory moves. Medvedev agreed to remove customs duties from exports of oil provided that the Belarusian side accepted the conditions of the Customs Union with Russia and Kazakhstan by January 1. Lukashenka agreed promptly to this request, which indicated his willingness to introduce the Russian ruble as the common currency once the Union comes into effect formally on January 1, 2012 (RIA Novosti, December 10). Lukashenka commented that the agreement would elevate relations with Russia to a new stage (www.telegraf.by, December 9).
The price for the latest Lukashenka victory could be a high one. Lukashenka will secure Moscow’s qualified support in the election (though Russian observers may offer some general objections to the way the campaign has been conducted) but he has effectively conceded economic sovereignty to his neighbor. Gas prices will rise in January 2011 from $185 to $225 per thousand cubic meters, and as candidate Viktar Tyarashchanka has noted, the average debt per Belarusian citizen is now $2,600 and the currency has depreciated against the US dollar by more than 4,000 times since 1994 (www.telegraf.by, November 25).
Whether Lukashenka needed to pay such a price is a moot point, but ultimately he responded to Moscow’s pressure to ensure his own immediate survival.
Belarusian Review, Vol. 22, No. 4
Copyright 2010 Belarusian Review
Source: Eurasia Daily Monitor Volume: 7 Issue: 225, December 16, 2010
Dr..David Marples
Printable Version Counter: 1118
Belarus' Forum
Belarusian Culture
Belarusians Abroad
Chronicle of Events
Legacy of Chernobyl
- Culture & Society
- Freedom of Religion
- Independent Media
- Belarus & the World
- Human Rights
- Politics - Opposition
- Politics - Regime
The Center for Belarusian Studies (23.01.2011)
Declaration by the Coordinating Committee of Belarusians of Canada (23.01.2011)
Blood and Special Operations in Belarus Politics (23.01.2011)
Belarus: That’s enough democracy (23.01.2011)
Wrong Carrot, Wrong Stick (23.01.2011)
Statement By the Rada of The Belarusian Democratic Republic in Exile
28th December 2010 (23.01.2011)
STATEMENT (23.01.2011)
BNR Rada Rejects Election Results (23.01.2011)
A backfire in Belarus (23.01.2011)
Demonstration in London (23.01.2011)
StatementBy the Office for a Democratic Belarus (Brussels)
And the Belarusian Institute for Strategic Studies (23.01.2011)
A Nasty Surprise in Belarus (23.01.2011)
In Belarus, a Slide Toward Eastern Aggression (23.01.2011)
Belarus Police Arrest Opposition Leaders (23.01.2011)
No Business as Usual (23.01.2011)
Belarusian Weapons Exports:A Possible Source of Laundered Funds? (15914)
The Exchange Rate Policy for Belarus (15758)
Kitabs, the Unique Phenomenon of the Belarusian Language in "Encyclopedia of the Belarusian language" (15700)
Belarusica at the AAASS Convention (14717)
Vaclav Havel - in Defense of Jakub Kolas Humanities Lyceum (14654)
Putin Doctrine: Immediate Threat to Belarus (14248)
Belarusian Currency: Problems and Perspectives (14115)
Is Lukashenka Winning Back Hearts and Minds? (13757)
DZIADY and Kurapaty Unite the Opposition (13422)
Belarus Represented in the World Book Fair (13403)
Minsk Wants Compensation from Moscow for Abandoning National Currency (13402)
Opposition Forms New Pro-European Alliance (13240)
The Sad State of Agriculture (12882)
EU Aid Misuse Fears (12390)
The Third Wave, or the regimes current tactics in dealing with independent NGOs (11967)
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Tue Apr 30, 2002 - An update - finally!
Ok, so here we are, sitting at an internet cafe in Hope, BC. Yesterday was a trying day, about 30 minutes after starting out I noticed my back tire was a little flat. Replaced the tube (lots of fun, since that required unpacking the bike, and repacking afterwards). Picked out a few pieces of rock and glass that were embedded in the tire, probably should do that each night. We got going at a nice clip for awhile, a few hills, but nothing major. Just after lunch, Jeff managed to get a flat (Just after saying that carrying 3 spare tubes was probably overkill, heh). We got going again, and not 2 minutes later we hear a loud *bang*. That was Jeff's tube bursting. I can still hear my ear ringing it was so damn loud. Ended up biking about 55km or so into Hope.
We're taking a rest day today, figured our knees and legs could use the rest. There's not much in the way of semi-large cities between here and Kamloops.
Once I have a bit more time I'll write a more detailed update :-)
Wed May 1, 2002 - Hope -> Lytton BC: 118km
Well, after a nice rest day in Hope, we felt like we could tackle a mountain, and so we did! (well, almost). Biking up the Frasier canyon is a beautiful ride, although some of the climbs are killers. Most of them we had to walk up, when you're carrying 60 pounds of gear, biking uphill for 2 or 3kms is rather difficult!
We left around 8am so it was refreshingly cold out. Nice biking temperature. It quickly became an extremely hot day though, so we took our time. Arrived in Lytton around 6pm. Today was the tunnel day, where we biked through 7 tunnels varrying from 100 metres to the longest which is around 700 metres long. There's no shoulder in the tunnels, so one option is to walk your bike along the pedestrian walkway, but that's about 3 feet wide which makes it really difficult to walk your bike. We opted to bike through most of them, and were a little worried about the longest tunnel where we wouldn't be able to see the other side to wait for a break in the traffic. As luck would have it, there was maintence work being done on the tunnels, so only one lane of traffic was being let through. We pulled up to the front of a huge line of traffic waiting at the mouth of the tunnel, and as soon as we arrived, it was the north bound lanes turn to go through! So we managed to get a head of most of the traffic going through the tunnel.
The traffic on the Trans-Canada is pretty light on this section, and the trucks are usually pretty nice to us, moving over to the oncoming lane if they can to give us more room. Luckily there's usually a lot of shoulder to ride on. At the end of the day we were pretty tired, so it was nice to set up the tent at a BC Parks campground, cook dinner (Stew, PBJ sandwhiches) and relax. The BC Parks campground was pretty empty, so we had a nice quiet evening to get lots of sleep. It's up early tomorrow to head to Cache Creek!
Thu May 2, 2002 - Lytton -> Cache Creek BC: 80km
Up early again at 6am or so, the mornings keep getting colder. Brr! The change of scenery over the span of about 20 kms is amazing, we're now entering a desert like area, very stark and barren in sections. I had no idea BC was like this!
I hit a piece of wood yesterday (was paying too much attention to the scenery, and not the road in front of me, oops!), so my back wheel needs truing. Hopefully there will be a bike store of a decent size in Kamloops. Other than that, the bikes have been amazing.
A few steep climbs today, which we once again walked up. We were up to 520 metres above sea level at one point. It's really tough to tell if you're biking uphill sometimes, since there's no good point of reference. We'll be biking along thinking it's flat and wondering why it takes so much effort! When you look back you realize that you've really been going up a slight incline all the way. There's almost no flat sections at all, either you're climbing, or going downhill. The Trans-Canada is cut into the side of the mountains at some point, a little scary! We thought we were going to get rained on, the clouds to the south looked pretty menacing, but we seem to have outrun them for now. It doesn't look like it rains much in this part of BC anyhow.
We got into Cache Creek fairly early around 2pm. There was a nice tail wind all day which helped is along, really ups the average speed quite a bit! Just a little while after setting up the tent, we look outside and see massive storm clouds rolling in. The wind was insane, almost thought the tent would be blown away! The temperature dropped about 20 degrees in a short amount of time. Well, I guess that's what you get for being in the mountains in early May!
We're off to Kamloops tomorrow, where we'll take another rest day. Getting used to carrying all our gear is taking awhile, but I think we're almost there.
Fri May 3, 2002 - Cache Creek -> Kamloops BC: 80km
Heh, they call this section of BC plateau, yeah right! There's more hills here then in the mountains! Climbed up to over 700 metres at times. We walked up more hills today then the previous two days combined! Overall a pretty good ride though. The air is slightly thinner at 700 metres then we're used to, took us awhile to figure that out, and to breath more deeply. We were wondering why we were having such a tough time of it!
We stopped for much needed food at Savona, as luck would have it there was a nice bakery so we had some sort of apple cinnamon pastry which was incredibly good! Speaking of food, almost anything tastes really good when you're hungry! The canned stew we've been eating tastes like the best stew I've ever had!
Quite a few cars (and some trucks) have been honking at us, makes us wonder if they have seen us biking on a previous day. It's a little unnerving though when a car honks right beside you, makes you lose a bit of concentration especially when you're biking uphill!
We've gone through a jar of peanut butter, and a jar of jam since Hope, so it's time to stock up again. Usually we eat about a loaf of bread a day. Plus Power bars, apples, or some sort of fruit. Subway has been a good place to eat as well :-)
The rest day in Kamloops will be much needed, it took us much longer to get anywhere today, muscles are getting a little tired :-) So far we've biked about 560kms, not bad at all!!
Sat May 4, 2002 - Kamloops - Ahhh, Rest Day!!
We're enjoying a day off in Kamloops, give our leg muscles a much needed break before the Rocky Mountain passes. Have to get my rear bike tire trued as well, and it's in the shop as I type. It was slightly bent due to that silly piece of wood on the road that I accidently biked over a few days back. It's probably a good idea to get the bikes checked out before the real mountains begin as well.
There's new pictures up as well, so have a look! Some of them turned out really good! Probably won't be able to update the site for awhile after this, the next major town is Revelstoke, and I doubt they'll have an internet cafe, so the next place is probably Calgary. Talk to y'all then! :-)
Wed May 15, 2002 - Maple Creek - Sask.
Only biked about 93km today. It was a little rainy in the morning, quite cold. Winds are still against us, we keep hoping they will change! The campground was a long way south of the Trans-Canada, and it was too late to continue on to the next town, so we found a cheap, run-down motel on the highway. At least we can catch up on the news tonight! Aparently there's a 'boil-water' advisory, so we'll have to buy some bottled water for tomorrow. (When we got to Swift Current, there really wasn't a 'boil-water' advisory, so we think the motel was just trying to sell bottled water).
The praries are quite a bit more hilly then I expected out here, although it will get flatter towards Winnipeg. Some parts are quite nice, some other parts are really dry, and they need rain badly.
Thu May 16, 2002 - Swift Current - Saskatchewan
Woke up to strong winds again, considering we have a 129km ride ahead of us, it could be a long day! After about 70km we started drafting each other, should have done this a long time ago! The first 70k were pretty hard, but the last 50k went by much much faster. Met up with the 2 cyclists we met earlier, they were a few minutes behind us. We'll probably meet them again at the camp ground in Swift Current. I think we pushed it a little too hard for the last 2 hours though, one of the muscles on my left leg is pretty sore. Nothing a rest day can't fix though!
Fri May 17, 2002 - Rest day in Swift Current - Saskatchewan
We haven't been near internet in so long, and we have only a short time at the library here, which means not much of an update. We'll update the site at the next chance we get hopefully.
Everything has been going well for the most part, the weather has been fairly cooperative, although we had a bit of rain a few days ago. The wind however is really slowing us down. Usually it blows from the west across the praries, but we've only seen crosswinds, or east winds so far. Despite that, we made it from Calgary to here in 4 days, which is about 520 kilometres. Jeff broke another spoke, and we don't have the tool to take the gears off, so we were hoping there'd be a bike shop here. Heh, yeah right, this is a pure ghetto town. The only internet cafe was shut down awhile ago, since the old folk didn't like it. There's no book stores either. Anyways, we're heading off to Moose Jaw tomorrow, about 170k away, lets hope the wind changes a bit!
Sat May 18, 2002 - Herbert Sask.
Well, we only biked 45km today, the wind was incredible! It's been blowing from the South East for quite a few days now, slowing us down quite a bit. We were pretty tired when we rolled into Herbert around 3:30pm, so we had to do some serious thinking. At this pace we wouldn't make our graduation on June 15th, so we'd have to take a bus from wherever we were at the time. The second option was to take the bus from Herbert to Winnipeg, skip about half the praries (which aren't incredibly interesting along the Trans-Canada highway anyway), which would give us more time in Northern Ontario. After much discussion, we decided to take the bus to Winnipeg. It seemed easy enough, but it proved to be quite the adventure! We intended to catch the 5:00pm bus, not realizing that our bikes would need to be in boxes before Greyhound would carry them. So missed that bus. Where does one find bike boxes in the middle of nowhere in Saskatchewan? At the gas station of course! The guys at the Co-op gas station were incredibly helpful, they managed to scrouge up some boxes which we could tape together to make a decent size bike box (Yay Duct tape!). The final result looks pretty interesting, we took a few pictures which should be up by now. The gas station even let us pitch our tent in the back to wait for the 4:15am bus. So off we went, made some dinner, and got a bit of sleep. Woke up at 3am to pack everything up. 4:15 rolls around, no bus. 4:30 - no bus. By 4:45 we were getting pretty cold, and pretty annoyed at Greyhound. Turns out somewhere they made a mistake, and there was no 4:14am bus. Oh well, what can ya do! At this point, we didn't want to set the tent back up, so we layed down the ground sheet, unrolled our sleeping bags, and slept under the stars! It was quite an experience. Thankfully, the 8:37am bus did show up, and we're on our way to Winnipeg!!
Tue May 21, 2002 - Winnipeg, woohoo!
We're here! Well, we've been here for 2 days now, getting some rest. Winnipeg is a really nice city! We're staying with my relatives, Catherine and Sid, who have been amazing! Had the whole tour of the city yesterday, and we've been doing some admistrative stuff today (fix bike, update website, etc).
Tomorrow we're heading out again, all of Northern Ontario is before us, and we're really looking forward to it! It'll be nice to see some hills again (never thought I'd hear myself say that!). We expect around 2 weeks to the Soo from here. Take a look at the pictures, some of them are pretty good! We'll be putting descriptions on all the pictures after the trip is over, we have them all written down, but no time to type them in right now.
Tue Jun 4, 2002 - Sault. Ste. Marie
While Jeff has been busy writing, I've been busy socializing, so sorry for the lack up updates!! When I get back to Pickering I'll have lots of time to write everything down. For now Jeff's journal will have to do . We're heading out tomorrow for Iron Bridge, taking the Manitoulin island route to southern Ontario.
After a rest day here we're feeling quite a bit more rested. We've biked 1400km from Winnipeg with only 1 rest day in between, so it was quite the ride! Only a week or so until we're back home, it'll be nice to sleep in a real bed again! Biking through northern Ontario wasn't nearly as bad as I imagined, although some of the hills were defintely challenging. The scenery is wonderful, especially through Lake Superior provincial park. Saw quite a few moose through that area, something I hadn't seen before.
This is the first major city where we haven't had to visit a bike shop. The bikes are holding up extremely well, and we're quite happy with them. We're especially happy with the tires, I haven't had a flat since before Calgary, and Jeff hasn't had one since before Hope BC. That's awesome for road tires!
Look back for updates in a week and a half or so! I promise I'll make time to get everything written down
Sun Jun 23, 2002 - The wheels on the bike go round and round....
Yes, Jeff and I are heading out bright and early tomorrow morning. We're well fed and rested after our 2 week vacation, so we're looking forward to getting back into the swing of things!!
These past 2 weeks have been pretty busy, visited Waterloo for a bit, saw some friends from Pickering that I hadn't seen in ages, and somewhere in there I graduated. Whew! All in all a fun time. But it'll be nice to get back on the road for a couple more weeks! We'll keep you posted on all our adventures, so stay tuned!!
Thu Jun 27, 2002 - Ottawa (167km)
Our longest day yet! We left Virgina's around 8:15 or so, hoping to avoid the heat of the day. Environment Canada was calling for Thunderstorms later in the afternoon, and while we welcome the rain to cool us off, lightning is not something we want be around. Many rest stops were needed to cool off, and to refil our water supply. Today was the first day I actually need to use the water in my canteen to refill my water bottles.
We stopped around noon for half an hour or so, but decided to keep biking. Not soon after we found some shade beside a church, so we stopped for some PBJ sandwhiches. While we were stopped however, a thunderstorm had sneaked up behind us, so we had to get moving! The wind for once was behind us, pushing us along at 35km/h. Amazing! We got rained on a bit, which was even better. We kept that pace up for about 20k or so, managing to get to Smith Falls ahead of the storm. Hid out in the local Tim Hortons for the storm to pass, which it did quite quickly. By this point it was about 3 o'clock, and we were just over halfway to Ottawa.
That last stretch of the ride was grueling, we're still not back into the shape we were before arriving in Toronto, but the thought of an air-conditioned apartment kept us going . Just outside of Smith Falls, a dog decided to make our day a little more interesting. It came out of nowhere, snarling and growling. I didn't see it at first, since most dogs bark well before they get near you. This dog was intent on biting our legs though, and I had to remove my foot from the toe-clip so I could kick it if it came any nearer (at that point it was running alongside about 3 feet away). We quickly outran it though, leaving us with a rush of adrenaline which lasted about 10k. Too bad I didn't get a chance to use the bear spray on it .
After that we had an uneventful finish to the day, traffic was fairly light except for a few sections nearer towards Ottawa.
Route: Hwy 15 to Smith Falls, then the 4, to the 6 to the 73 up to Ottawa.
Sat Jul 6, 2002 - St. Barthelemy (128k)
We got off to a slow start today, taking a 10k dead end. Well, we thought we could take a highway that cyclists were not allowed on, oops! So we backtracked and followed the Tour du Canada route once again. It seems every time we detour from their route, something goes wrong, so we've learned to stick to it .
We followed the Richelieu river almost all the way from Montreal to Sorel which is right on the St. Lawrence river. The entire length of the Richelieu is lined with houses, and boats were out on the river enjoying the afternoon. We stopped for a break in a small town where an art exhibit was taking place. A little while later in another town, we had a "conversation" with another cyclist, although he seemed to be speaking Italian, and of course, we only speak english! We compared our bike computers to see how far we'd gone, he was around 3500k, so pretty impressive! We crossed the St. Lawrence on a ferry at Sorel, picked up some dinner in the town immediatly across the river, and made our way to the campground. What we didn't realize was the campground was 8k off the highway! It turned out to be a decent campground, more of a resort really. They had lots of "social" events for familes, and so on. There was another cyclist staying at the campground, heading east towards Quebec as we are. We didn't catch her name though, so we arbitrarily named her Anne.
Sun Jul 7, 2002 - Portneuf (130k)
A pretty uneventful day. Anne (see previous entry) left an hour before us, but we caught up to her fairly quickly. A helpful west wind was pushing us along at about 30km/h. Arrived at another "resort" campground. The guy next to us insisted on playing his music quite loudly, and we were woken up by noisy children the next morning. It'll be nice to go to some quieter campgrounds!
Mon Jul 8, 2002 - Quebec (60k)
One of our shortest days yet, but there's too much smoke in the air to make it worth biking. You can barely see across the St. Lawrence! Combine that with some nasty allergies, and we didn't feel too good.
We're staying at Laval University residences, which is a welcome change over our tent. A good nights rest is in order! Quebec City has a fairly extensive bike path network, once again proving that Toronto really needs to work on their bike paths!
Tue Jul 9, 2002 - Quebec City
I woke up this morning quite refreshed! Both of us needed a good sleep after the past few days. Allergies combined with not so comfortable bumpy ground has prevented us from sleeping very well. On the agenda today was tour around Old Quebec, update the website, and do a bit of maintenance on our bikes.
Quebec is blanketed in smoke from the fires burning north of Montreal. At times visibility is a couple kilometers at most, you can barely see across the St. Lawrence river from the boardwalk in front of Chateau Frontenac. The smoke appeared a few days ago, which at first we thought was smog due to humidity. We didn't find out it was a fire until later on when reading a newspaper (which was in french, so we only had a rough idea of what was going on).
Wed Jul 10, 2002 - St. Jean - Port Joli (118k)
This was by far our strangest day yet! First off we had to figure out how to cross the St. Lawrence. With only 2 bridges, one of which we can bike on, you'd think it would be easy, right? Nope! It wasn't well marked, so it took us a while to figure out where to go. The bridge is huge steel monstrousity that doesn't feel safe at all. In fact the middle section has fallen out once before. The pedestrian walkway was just wide enough to allow for a single bike. Once on the south shore, we pointed our bikes east, and started cycling. We didn't get very far before we witnessed the first car accident of the trip, indirectly caused by us! Here's the scoop: We were biking through an intersection on a green light. An oncoming truck was making a left-hand turn across our path, but he ran a red light making that turn. Halfway through the intersection, he noticed us, and stopped. A few seconds later, a white Honda Civic heading the same direction as us plowed into the side of the truck. There wasn't much damage, the truck was unharmed, and the Civic had it's left headlight taken out. What we can't figure out was how the Civic didn't notice the truck sitting in the middle of the intersection! Obviously the driver wasn't paying attention to the road in front of him. We ate a bit of lunch while we waited for the police to show up. When the officer finally got around to talking to us, our statement consisted of "The light was green". By this time it was 2pm, and we're barely out of Quebec City, so we had to get a move on!
About 4:30 we met a guy walking from Ottawa to the East coast, so we chatted to him for a bit. Jeff just checked his website though, and he appears to be way ahead of us, so he must have cheated a bit and hitched a ride somewhere.
Arrived at the campground quite late, just as it was getting dark. This is by far the most beautiful camp site yet! It's right on the St. Lawrence, which at this point is probably at least 5 or 10k across. Very nice view of the mountains on the other side.
Thu Jul 11, 2002 - Riviere-Du-Loup (95k)
Riviere-Du-Loup means River of the Wolf or so I'm told. Thankfully we didn't see any wolves though! This was an incredibly scenic day! The highway runs right along the St. Lawrence, giving us an awesome view of the mountains on the other side. This side of the river is gentle rolling hills, mostly farmland in-between forests. The weather couldn't be any better, a mix of sun and cloud, and a perfect temperature for biking.
We met up with 2 other pairs of cyclists at the campground, so it was nice to chat for a bit, and compare stories. One of the pair was from Montreal, and the other was from somewhere in North Eastern Quebec (can't rememeber exactly where at the moment). We saw another pair of cyclists earlier in the day, but they didn't stay at this campground.
Fri Jul 12, 2002 - Saybec (190k)
Yes, that's really a 190 up there! Our longest day yet by far. We had a strong west wind to help us along, crusing usually over 30kph. I broke my max speed for the trip, hitting 77kph down a hill. Biking at that speed is incredible! The roads however, leave much to be desired. Sometimes there's a shoulder to ride on, more often there's not. A lot of the time the road is riddled with pot-holes, and we have no choice but to ride through them. My back rim took quite a beating, and needs to be trued badly, or else rebuilt with new spokes.
We saw the pair of cyclists from yesterday (the ones who didn't stay at our campground). We caught up to them, passed them, and they caught up to us a couple times when we stopped for a break. Nothing like some friendly competition to keep us going <grin>. We arrived in Mont. Joli in mid-afternoon, not bad for 140k. I was quite prepared to call it a day, but Jeff had energy to burn, and so did I as it turned out, so we continued on. A few km's down the road, we looked back to see huge black rain clouds blowing in from the north. The race was on! Unfortunately we lost, and got rather soaked. The terrain changed from almost perfectly flat to extremely hilly, making the last 50k to Saybec quite difficult. Our tent site for the night was quite silly. It was a patch of freshly sodded grass so small it would barely fit our tent. Keep in mind that we have a 2 person tent which isn't large to begin with! So we weren't too happy about that, not to mention being wet and tired. Overall it was a really good day though, I doubt we'll break 190k on the rest of the trip!
Sat Jul 13, 2002 - Campbellton - NB (136k)
Our seventh province! Whoohoo! Well, after cycling 190k yesterday, we turned around and did another 130k today! Our muscles are complaining rather loudly however, and some rest is defintely in order. It rained pretty much all day, and yet it was sunny for most of the day as well, go figure. I think a rain cloud had nothing better to do except follow us around all day <grin>. We followed the Matepedia river and the landscape is quite impressive. Just like the mountains of BC, but on a much smaller scale.
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BitCharts.io
Bitcharts Issues ‘Breakout Watch’ on Bitcoin at $11,800
‘BitChart’ Adds New Features and Coins to its Bitcoin Tracking App
AppSwarm (SWRM) Announces Launch of Bitcoin Tracking App ‘Bitchart’
Home / ICO News / BitChart / AppSwarm (SWRM) Announces Launch of Bitcoin Tracking App ‘Bitchart’
Tusla, OK — January 10, 2018 — InvestorsHub NewsWire — AppSwarm, Inc. (OTC:SWRM), a technology company specializing in the accelerated development and publishing of mobile apps, announces the beta launch of its new Bitcoin tracking app ‘Bitchart’.
Bitchart Mobile App
BitChart is a sleek and interactive way to visualize the bitcoin price index through your smartphone. Always be up to date on the current spot price and the daily change. Utilize real-time alerts to keep an eye on price shifts. Analyze historical trends with the swipe of a finger. Easy access to the current spot price with the BitChart Notification Center widget. Support for USD and EUR across the most popular exchanges.
Still in the Beta stage, we plan to incorporate news coverage of the cryptocurrency market, with research analysis, interviews, and real-time chat feature with other cryptocurrency traders from around the globe.
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Ron Brewer, CEO of AppSwarm, Inc. commented, “We are very excited about the beta launch of our new Bitchart app, and think going forward we have the potential to build a unique investors hub for crypto investors, while also helping bring some additional transparency to this rapidly growing market. Through our years of mobile development of games and different business applications, we look forward to adding a number of new features to our Bitchart app through the rest of 2018, and will be providing updates on this in the very near future.”
The beta version of Bitchart can now be downloaded on the front page of our new site at http://www.bitcharts.io
About APPSWARM:
AppSwarm is a technology company specializing in the accelerated development and publishing of mobile apps and other software platforms for gaming and business applications and seeks to acquire symmetric business opportunities. AppSwarm partners with and assists other development firms in technology development, business management, and funding needs.
For more information, visit us at www.app-swarm.com, or follow us
on www.facebook.com/AppSwarm or Twitter twitter.com/AppSwarm
Forward-Looking Statements:
“Safe Harbor” statement under the Private Securities Litigation Reform Act of 1995: This press release may contain forward-looking statements that are subject to risk and uncertainties including, but not limited to, the impact of competitive products, product demand, market acceptance risks, fluctuations in operating results, political risk and other risks detailed from time to time in the Company’s filings with OTCMarkets.com and as required to the Securities and Exchange Commission. These risks could cause SWRM’s actual results to differ materially from those expressed in any forward-looking statements made by, or on behalf of, the Company.
Investor and Media Contacts:
AppSwarm, Inc.
info@app-swarm.com
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Archive for activities
Seven things you should do before moving to NYC
If you are preparing to embark on the SIPA adventure this fall to spend the next two years of your life in New York City, here is some advice on what you should definitely try and do before boarding that plane.
I know, you are permanently moving to New York for at least the next two years, and might feel tempted to bring along those pants that you may someday want to wear, or that old lamp you love. But New York is crowded, and space is not precisely a surplus commodity. Whether you live in Columbia housing or on your own, most likely you will have a small room, and an even smaller closet.
Read a novel
For those of you who like reading novels; this will probably be one of the first pleasures cruelly swiped away from your life by grad school. There is so much to read for every class, that reading a novel simply becomes a luxury that a SIPA student cannot afford. So use this summer to indulge in those fictitious adventures, as they will be deeply missed.
New York is an amazing city, but it can also be amazingly expensive. If you want to have an occasional dinner at a nice restaurant, go to a concert, or take a taxi to avoid a 2-hour ride back from Brooklyn on a Saturday night, you should try and save some money to help you enjoy the city more.
Go see nature
You’ve heard the song. New York is indeed a unique concrete jungle where dreams are made of. But as fascinating as skyscrapers can be, they can also be overwhelming. So make sure you get a good dose of wild mountains and blue oceans before you head this way.
Get some rest
SIPA will be lots of fun, but also lots of work. You will have endless nights in our basement library, for which you will need plenty of energy to help you cope. So get some serious sleep and rest before going back to school.
Let go of your prejudices
If you are coming to SIPA, you are probably already on track, but it doesn’t hurt to think about this once in a while. New York is a truly diverse place, and that is a central part of its magic. So open your mind and be ready to learn from other worldviews, cultures, careers and human beings. The more prepared you are to learning new things, the better your experience will be.
Be ready to be merry
Grad school, for most of us, happens once. Chances are this will be the last time in your life to be a student at a formal academic institution. Be consciously grateful for the endless opportunities, experiences and freedom the next two years will give you.
Categories : Admissions, Application Tips, Columbia University, SIPA
Tags : activities, events, moving to NYC, new york, preparation, summer
Journal of International Affairs
Posted by Matt Clemons
· Thursday, February 3rd, 2011
The following entry was contributed by Homa Hassan. Homa is a first-year student at SIPA and you can read her biography here. There are plenty of extracurricular opportunities for SIPA students to get involved in and one such opportunity is with the Journal of International Affairs. Homa elaborates on her experience as both an Editorial and Digital Assistant.
SIPA students are always looking for ways to get involved on campus. There is a mix of activities to take part in – there are the organizations that have to do with your particular studies, the ones that deal with your personal interests, the ones with tradition and prestige, and the ones that introduce you to new fields and new friends.
One of the projects that links new students to old and all students to alumni and global leaders is Columbia University’s Journal of International Affairs. Two volumes are produced each year comprised of essays, interviews, and book reviews written by prominent scholars of foreign policy and international affairs. The Andrew Wellington Cordier Essay contest also gives students the opportunity to have their original works published. The theme of each volume is decided on by SIPA students on the JIA Board. Our latest theme was on Innovating Development, with essays ranging from nuclear power and sustainable development to climate change technology and gender revolutions.
There are a number of different ways to get involved with the Journal as a first year student, including serving as an Editorial Assistant, Production Assistant, and/or a Digital Assistant. No experience is necessary and training is provided for each of the positions at the beginning of the semester. I served as both an Editorial Assistant and a Digital Assistant. Editorial Assistants have a demanding job. We don’t just copy edit, which of course is important, but the bigger task is to fact-check. Editorial Assistants are the first defense on the line protecting the publication’s reputation.
To begin with, each of us was given a brief summary of all of the articles and asked to rank them in the order of our interest. A few days later, we were assigned 2-3 pages in one of the essays and paired with a Lead Editor who oversaw our work. For our assigned pages, every line had to be checked and re-checked and verified by two or more sources (online or in print) to ensure the information is accurate and well-supported by substantial evidence. Any links in the article were tracked and also verified. The work is intense, but one of the most important processes of publishing the Journal.
The Production Assistants work with the layout of the publication, placing the content onto the pages, formatting and designing them to be aesthetically appealing, and finalizing the essays before they are sent out for production. When the publication is finally out, the Board hosts something akin to a release party. This year, it was a nice break from studying for final exams to listen to the speakers’ panel and join the celebrations at the subsequent reception with food, drinks, and live jazz music. The JIA Board also hosted an end-of-semester happy hour to thank all of the SIPA students who helped out.
Digital Assistants continue to work throughout the year. One of the large projects JIA has right now is uploading all previous issues of the Journal to the JIA website. This means sorting through PDF versions of old issues and uploading the content to the website so that they are accessible to the general public. The nice part about being a Digital Assistant was that I could go into the publication room between classes or after work and get my work done independently. Having worked on the production and editorial side of journalism in the past, I welcomed learning new skills by working on the digital side of things. I also now know the ins and outs of the publication online (great resource for classes!).
Working for the Journal of International Affairs has been challenging, but one of the most rewarding ways to get involved at SIPA. Its fluid structure allows you to try new things and advance with commitment. And years from now, your work is still an integral part of SIPA’s library. Working on the Journal allows you to hit the ground running and seamlessly integrates you into life at SIPA.
Categories : Meet Seeples, News, SIPA
Tags : activities, journal, publishing, research
A Day in the Shoes of a SIPA Student
· Tuesday, December 14th, 2010
What is a “typical” day like for a SIPA student? Carrie Dorn offers her perspective . . .
When I first informed my friends about my acceptance to SIPA, they gave me a lot of unsolicited advice and some of it turned out to be very useful. I was advised to pack in all of the rest and relaxation that I could get before the school year started by catching up with acquaintances and family members, organizing my house, reading some non-fiction, seeing the newest movies, and taking a vacation. Being the type of person who has always had a busy schedule, I didn’t imagine that the life of a student would be much different from my work life.
But, now that I am at SIPA, I pass this wisdom on to you. Life at SIPA is busier and more exhilarating than you can anticipate. Between classes there are lectures attend, cover letters to write, and networking to be done. At night there are socials to go to, documentaries to watch, research to gather, and group presentations to practice. The days of a student are long, the homework never-ending, and always learning opportunities not to be missed.
Here is a snapshot of what my day looked like on a fairly typical Tuesday in mid-November . . .
Wake up at 6 am and brew some coffee. Feeling awake!
Read and respond to emails and proofread a paper proposal to be handed in tomorrow. Running late!
Power walk to my 9 am class, Advanced Generalist Practice and Programming at the School of Social Work. Today’s topic is integrating qualitative and quantitative data for comprehensive asset-needs assessments. Meet with my group after class to discuss interviews and edits for our project, determining the strengths and needs of individuals reentering the East Harlem community after incarceration. So much to be done!
Report to SIPA’s Office of Admissions and Financial Aid for work at 11:15 am. Talk to prospective students on the phone and in person about the impressive work that they’re doing and their future career goals. Extraordinary applicants!
At 2 pm, meet with a fellow student and plan for our upcoming conversations with administrators about the inter-school coordination of dual degree programs. Advocating for improved processes!
Stop by the Office of Career Services to schedule an appointment to review my resume with a counselor. Find out the dates and times of upcoming New York City Career Panel sessions. Filling up my planner!
Report back to the Office of Admissions for the afternoon hours to assist in filing and answering questions about the admissions process by email.
Go to the 6th floor computer lab to print out materials for tomorrow’s classes and chat with friends.
At 8pm go to the Brazil Trip information session to find out about SIPA’s Spring Break Trip to São Paulo and Rio de Janeiro. Pizza dinner provided!
Get home around 9:30 pm, finish up homework and plan for tomorrow. Jump into my comfy bed at midnight and fall asleep. Dream until the alarm goes off tomorrow morning!
Categories : Columbia University, Meet Seeples
Tags : activities, classes, student life
· Thursday, December 2nd, 2010
Below is some evidence of the choices that SIPA students must sometimes make when it comes to how to spend their time. There always seems to be something going on at SIPA or on our campus that would be interesting to attend.
Gender-Based Violence in the Congo
International Affairs Building, Room 1501
Panel Discussion with Dr. Les Roberts, Professor of Epidemiology at Columbia University Mailman School of Public Health; Paula Donovan, Co-Founder of AIDS-Free World; Dr. Susan Bartels, Co-Head of the Harvard Humanitarian Initiative; and Lisa Jackson, Writer and Director of the film “The Greatest Silence: Rape in the Congo.”
Debate: Nuclear Energy and Climate Change
Columbia Law School, Jerome Greene Hall, Room 106
Earth Institute
Debate with Robert Alvarez, Senior Scholar, Institute for Policy Studies, former Senior Policy Advisor to the Secretary of Energy; Peter Bradford, Adjunct Professor, Vermont Law School, former Commissioner, Nuclear Regulatory Commission, former Chair, New York and Maine utility regulatory commissions; Barton Cowan, Visiting Professor, West Virginia University College of Law, of counsel, Eckert Seamans Cherin & Mellott, LLC; Susan Eisenhower, Member, Blue Ribbon Commission for America’s Nuclear Future, Chair Emeritus, Eisenhower Institute; Michael Gerrard, Andrew Sabin Professor of Professional Practice, Director, Columbia Center for Climate Change Law
Kazakhstan’s Refugee Crisis: Violence, Hunger and the Transformation of Broader Central Asia, 1930-1933
Harriman Institute
Lecture with Sarah Cameron , Post-Doctoral Fellow, Yale University
Japan Circa 1959 – The High-Growth Economy and the Social Effects of Television
International Affairs Building, Room 918
Weatherhead East Asian Institute
Brown Bag Lecture with Yoshikuni Igarashi, Associate Professor of History, Vanderbilt University
International Affairs Building
Lecture with Sarah Cameron, Post-Doctoral Fellow, Yale
DevInfo Training
New Media Task Force
Workshop with Christina J. Irene, a representative from the joint UNICEF/DevInfo programme, along with the Fall 2010 DevInfo Interns, will present an introduction to the DevInfo data management system.
Brown Bag with Amb. Paul R. Seger, Permanent Repepresentative of Switzerland to the UN
International Organization Specialization
Brown Bag Lecture with Ambassador Paul R. Seger, Permanent Representative of Switzerland to the United Nations
How Not to Help
Institute for the Study of Human Rights
Discussion with Kate Cronin-Furman and Amanda Taub from “Wronging Rights”.
Lake Baikal, Siberia: Will Industrial Development Destroy the World’s Largest, Cleanest Lake?
Czech Foreign Policy After the Fall of Communism
Lecture with Jiri Paroubek
Perspectives on Political and Economic Dynamism in Northeast Asia- Challenges of China and North Korea
Columbia Univerity Morningside Campus International Affairs Building, Room 918
Center for Korean Research
Lecture with Ambassador Young-Mok Kim,Consul General of Republic of Korea to New York. No registration is required.
Leaders in Global Energy: Dr. Fatih Birol: Critical Factors Shaping the Future Global Energy Landscape
School of International and Public Affairs and Center for Energy, Marine Transportation and Public Policy
Lecture with Dr. Fatih Birol, Chief Economist, International Energy Agency
Tolerance Without Liberalism: Conflict and Coexistence in Twentieth-Century Indonesia
Center for the Study of Democracy, Toleration and Religion
Lecture with CDTR Visiting Fellow, Jeremy Menchik
My Perestroika
Film Screening and Discussion with Robin Hessman. To reserve tickets in advance please follow the link: www.ovationtix.com/trs/pe/8563295. Tickets will also be available at the box office in the Lerner Hall Lobby the day of the show.
Concert Series: Italian Harpsichord Music with Andrew Appel
The Italian Academy at Columbia, 1161 Amsterdam Avenue
The Italian Academy for Advanced Studies in America at Columbia University
Concert Series with harpsichordist Andrew Appel, violinist Krista Bennion Feeney, and cellist Loretta O’Sullivan, performing the music of Boccherini, Cimarosa, and Clementi
A Conversation with Adolfo Carrion, Regional Director of the U.S. Department of Housing and Urban Development
12:00 pm- 1:30 pm
Urban and Social Policy Concentration
Conversation with Adolfo Carrion, Regional Director of the U.S. Department of Housing and Urban Development
Migrations of Jewish-Hungarian Professionals through Germany to the United States, 1919-1945
East Central European Center
Discussion with Professor Tibor Frank, Eötvös Loránd University, Columbia University, regarding the impulses influencing a uniquely gifted generation of mostly Jewish Hungarian emigrants.
Biological Measures of the Standard of Living North and South of the Border –
Lecture: with Prof. Richard Steckel, Distinguished University Professor of Economics, Anthropology and History at Ohio State University.
When China Met Africa and The Colony
Studio X 180 Varick Street New York, NY 10014
Committee on Global Thought
Film screening / Discussion including two films that examine Chinese investment in Africa
Stories of Stigma, Stories of Strength: Ethnographic Oral History with Sanitation Workers in New York City
Schermerhorn, Room 754
Oral History Master of Arts Program
Lecture with Robin Nagle. She will present her ethnographic work for her forthcoming book Picking Up.
QMSS Seminar: Sexual Networks and HIV Transmission in a High-Prevalence Setting: Evidence from a Sociocentric Study
Hamilton Hall, Room 503
Institute for Social and Economic Research and Policy
Seminar with Stephane Helleringer, Mailman School of Public Health
Afghanistan: Prospects for Peace
Kellogg Center, International Affairs Building, Room 1501
Saltzman Institute of War and Peace Studies
Sixth Annual Arnold A. Saltzman Forum
From a Raindrop to a Stream Pebble to a Delta: Recent Research on Predictive Modeling
Seeley W. Mudd Building, Room 833
Lecture with Efi Foufoula-Georgiou, Distinguished McKnight University Professor, Director of the National Center for Earth-surface Dynamics, University of Minnesota
Asia in Africa: New Connections in Historical Perspective
Davis Auditorium, Schapiro Center
Discussion Panel with Howard French, Deborah Brautigam, Abdoulie Janneh, and Wang Hongyi
The International Criminal Court in Motion – An Analysis of its Seven Years of Activities and Perspectives with Dr. Luis Moreno-Ocampo, Prosecutor, International Criminal Court
International Affairs Building Room 1501
Center for International Conflict Resolution
Lecture with Dr. Luis Moreno-Ocampo, Prosecutor of the International Criminal Court (ICC). The talk will be followed by a discussion moderated by Mr. Jean-Marie Guéhenno, Director of the Center for International Conflict Resolution.
Toxica Simulation
CRWG and LASA
Simulation allowing participants to engage in a negotiation, observed by negotiation practitioners. Space is limited, RSVP required. Please email Toxica2010@gmail.com.
From Three-Legged to Two-Legged Races – The Emergence of Women’s Competitive Sports in Japan (1910s-20s)
Brown Bag Lecture with Robin Kietlinski, Adjunct Assistant Professor of History, Baruch College; Visiting Researcher, Weatherhead East Asian Institute.
Monday, December 6 – Distinguished Lecturer Series “Southern Buddhism: Tracing Later Buddhist Art in South India”
Knox Hall, Room 208
Southern Asian Institute
Distinguished Lecturer Series with John Guy, Metropolitan Museum of Art
Innovating for Development: A Thought Leadership Forum from the Journal of International Affairs
School of International and Public Affairs
Forum moderated by Steven Cohen, Executive Director, Earth Institute, about how innovation is driving the agenda for sustainable development, climate change, natural resource use and energy policy.
U.S. Rapprochement with Indonesia – From Problem State to Partner
Brown Bag Lecture with Ann Marie Murphy, Associate Professor, School of Diplomacy and International Relations, Seton Hall University; Adjunct Research Scholar, Weatherhead East Asian Institute.
Transforming Humiliation and Violent Conflict Workshop
Columbia University, Teachers College Campus, 525 West 120th Street, Grace Dodge Hall, Room 179
QMSS Seminar: Political Conditions for Diffusion? Anti-Corporate Movements and the Spread of Cooperatives in America Capitalism
Lecture with Marc Schneiberg, Queens College Department of Sociology
Post-Cancun Debriefing
The Columbia-Paris Alliance Program and the Sustainable Development Doctoral Society
Seminar on the climate change negotiations in Cancun, with Scott Barett, Lenfest-Earth Institute Professor of Natural Resource Economics and Laurence Tubiana, Alliance Visiting Professor at Columbia
From Wednesday, January 12, 2011 through Friday, January 14, 2011
SIPA Students Only: 35th Annual Washington, DC Career Conference
Office of Career Services, School of International and Public Affairs
35th Annual Washington, DC Career Conference, a three-day event consisting of 20 panels, employer site visits, networking reception and a day of informational interviews. For further information regarding this event, please contact Joe Musso at sipa.dc.conference@columbia.edu.
Categories : Academics, Columbia University, News, SIPA
Tags : activities, brown bag, events, lecture
Thursday Evenings at SIPA
· Thursday, December 10th, 2009
The following was contributed by Kristoffer Tangri, a second-year SIPA student from Germany pursuing a MIA degree with a concentration in International Security Policy.
Thursdays are popular for events and receptions at SIPA and sometimes it can be difficult to choose. Last Thursday was one of these days. After having listened to the insights of a respectable guest speaker from the International Peace Institute in my class on “Building Peace after Conflict”, I had a remarkable choice of public events and reception to attend at SIPA. Not always an easy choice.
Downstairs on the 4th floor, the auditorium was filling with students who were interested in hearing Noam Chomsky’s opinions on “The Unipolar Moment and the Culture of Imperialism”, while in the building next door, Robert C. Orr, Assistant Secretary-General for Planning and Policy Coordination in the Executive Office of the Secretary-General, was speaking on “The Secretary-General’s Agenda and The Challenges He Faces.”
Meanwhile, on the 15th floor, SIPA’s Energy Association was hosting a panel to discuss “the adaptation of large-scale renewable energy under a proposed cap and trade system” and on the same floor, Columbia’s Dirk Salomons was moderating a panel discussion about the international response to the ongoing crisis of childhood malnutrition with experts from the Doctors Without Borders.
After so much academic input, one feels the need to go out and socialize with fellow students – but where? Should I go to the UN Studies Christmas Party on the 9th floor, or drop by the Latin American Association’s reception? Maybe I go over to the B-School on Campus for a few hours of free drinks (paid for by the tuition of our wealthy Business School students). Later that evening, the Migration Working Group was having a fundraiser party down in the East Village, too.
One thing you can be sure of at SIPA. You will always have an amazing variety of public lectures and events to attend, both at SIPA and at the many departments around Columbia University and of course New York City itself, with the UN and Wall Street just around the corner. And in case you are worried about the living costs of New York, these events always come with free food and wine.
Categories : Admissions, Columbia University, News
Tags : activities, discussion, extracurricular, panel, speakers
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airBaltic to Launch Riga – Thessaloniki, Greece
22.01.2015 23:09 | Latvia
Photo: www.airbaltic.com
Author: Jānis Vanags
Source: www.airbaltic.com
airBaltic, the world’s most punctual airline, will launch a new route between Riga and Thessaloniki (Greece) this summer, on June 5, 2015. The new route will add another popular summer destination for Baltic travellers, as well as connecting travellers from the south with Riga, and beyond to destinations in the Baltics, Europe, Scandinavia, CIS and the Middle East.
Christophe Viatte, Chief Commercial Officer of airBaltic: “Thessaloniki is renowned for its vibrant cultural life, its ancient history which is visible on the facades of its impressive churches, monuments and buildings, while also being on the shores of the Aegean Sea. It is an ideal holiday destination for culture lovers and sunshine seekers alike. We are delighted to be adding this destination to our route map, broadening summer holiday options for travellers from the Baltic region, and strengthening direct links between northern and southern Europe.”
Flight frequency
Price *,
Price*,
Riga – Thessaloniki
2 flights weekly
* Lowest fare, including taxes, fees and service charges, on www.airBaltic.com, subject to availability
airBaltic will fly from Riga to Thessaloniki twice a week on Fridays and Sundays. Passengers will board a Boeing 737-300 aircraft for a flight that will last 2 hours and 55 minutes. The price of a one-way ticket will start at EUR 85, including airport fees and transaction costs. A full schedule of airBaltic flights can be found on the company's homepage at www.airBaltic.com.
airBaltic serves over 60 destinations from its home base in Riga, Latvia. From every one of these locations, airBaltic offers convenient connections via Riga to its network spanning Europe, Scandinavia, CIS and the Middle East.
airBaltic in brief:
AIR BALTIC CORPORATION (airBaltic) is a joint stock company that was established in 1995. Its primary shareholder is the Latvian state, which holds 99.8% of the stock. The airBaltic fleet consists of 25 aircraft – 13 Boeing 737 and 12 Bombardier Q400Next Gen. airBaltic offers nonstop flights from its hub in Riga, Latvia. airBaltic received the title of Airline of the Year 2009/2010 (Gold Award) from the European Regions Airline Association. airBaltic is the winner of Air Transport World Phoenix Award 2010 - a global recognition of excellence in restructuring business. airBaltic won a special Pacesetter Award 2010 from the Jury of the Budgies World Low Cost Airline Awards, recognizing the achievements of airBaltic as a hybrid airline. In 2012, airBaltic was ranked by Airlinetrends among the Top 10 airlines globally for innovations.
The event guide for the year 2020 has been released (17.01.2020)
How to fall in love with Riga in winter? (09.01.2020)
Motivate yourself to travel and buy a trip at «Balttour 2020!» (06.01.2020)
Four exhibitions from the Rothko Centre's collection (03.01.2020)
airBaltic Receives its 22nd Airbus A220-300 (27.12.2019)
airBaltic Offers Extra Flights During Christmas Holidays
airBaltic Airbus A220 Full-Flight Simulator Certified by CAA
The Business of Cargo Shipping is Rocked down by Waves Now
airBaltic in Canada Celebrates its 21st Airbus A220-300 in a New Livery
airBaltic Launches New Direct Routes from Tallinn and Vilnius
airBaltic Top Winter Holiday Destinations
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Review: August, Osage County
Dallas/Ft WorthReviewsTheater
August, cialis Osage County WaterTower Theatre, buy cialis Addison April 12 – April 22, viagra 2012.
August: Osage County, Tracy Letts’ powerful, punishing, acerbic comedy on the internal collapse of Western Civilization, received a formidable, well articulated production at the hands of Rene’ Moreno, directing at Water Tower Theatre. Pam Dougherty (as matriarch Violet Weston) leads a well-orchestrated, thirteen-member ensemble through a three hour joyride of frantic, grotesque, destructive, hysterical, rambunctious melodrama. The Westons are a white bourgeoisie clan with a Native American housekeeper, temporarily inhabiting a large house outside Pawhuska, Oklahoma. They have gathered for the funeral of patriarch Beverly Weston, a somewhat celebrated, alcoholic poet. At the opening of August, we see him interviewing Johnna, the previously mentioned domestic. He explains that he and Violet have an “arrangement” (meaning they must abuse substances to endure one another) just before he disappears. Like many tumultuous romances, apparently Beverly and Violet get along better in different rooms.
It seems pretty clear that the character of Johnna has been placed in August, for the sake of contrast. Violet, in a typical moment of nasty “truthfulness” points out that Johnna’s getting paid, when someone thanks her for cooking the meal. As if remuneration precludes appreciation. Johnna remains tranquil and resourceful even as family members try to navigate the chaos of their dysfunctional lives. Violet treats her as interloper, despite the fact that she undeniably makes things easier for all of them, and stays out of the way. When Letts raises issues like the cultural differences between Indians and Anglos, the debatable ethics of acquisition by conquest, the consumption of meat and the use of hallucinogenics, well, comparisons seem inevitable. She may not have a lot to say or more than one room to call her own, but Johnna is obviously more evolved than any of them. In a moment of crisis it is she that Violet seeks out.
You have to admire the way Letts has depicted the Westons, blameless and yet somehow tainted and pathetic. Lost and unstable, yet normal. Violet and her sister, Mattie Fae (and her husband and son) three grown daughters : one with a daughter and estranged husband, another with a fiancé, another single. Each one has their issues (as all of us do) but for some reason, when they’re all under one roof, eruptions flood like Krakatoa. Violet is addicted to prescription pills, teenaged granddaughter Jean smokes pot, Ivy Weston has fallen in love with first cousin, Little Charles. It’s not exactly like those lofty dynasties in the Greek tragedies, where crimes that would make the Manson family cringe, bring curses from Mount Olympus. But we do get the impression Letts is asking us to take a step or two back, and a long, sharp look at the failure of our culture to even gravitate towards spiritual enlightenment.
— Christopher Soden
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Home > People > Armed Forces Veterans > Roll of Honour -WW1 > Alfred John Westerman >
Ashwell Records
Renovation 2019
Friends of Ashwell Museum
Ashwell at Home
Jubilee Photo Archive
Alfred John Westerman
WWI Roll of Honour
By Lynette Wray
Military History:
Regiment: Royal Field Artillery
Rank: Driver
Regimental Number: 895730
Date of Enlisting: 1.11.1915
Date of death: 5.3.1950
Cemetery/ Memorial:
UK Cemetery/Memorial: St Mary’s Church Ashwell Roll of Honour
Alfred enlisted with the Royal Field Artillery Territorial Force, Regimental Number T895730, on 30th October 1915 at Hertford, Hertfordshire. He was posted to the 6C Reserve Brigade, Regimental Number 895730, on 26th February 1917 he was transferred to the 52nd Battery and on 25th May 1917 he was embarked to the Western Front, France and Flanders as a driver with the Royal Field Artillery. Alfred was demobbed on 31st August 1919.
Alfred survived the War and died at Shortstown, Bedfordshire on 5th March 1950.
Alfred was awarded the British War Medal and the Victory Medal.
He is commemorated on the St Mary’s Church Ashwell Roll of Honour.
Family History:
Date of Birth: 11.12.1896
Birth Place: Ashwell, Hertfordshire
Residence: Bedford, Bedfordshire
Marriage: Single
Alfred was born at Ashwell, Hertfordshire on 11th December 1896, the second son of Arthur and Georgina (nee Gray) Westerman. He was a pupil at the Merchant Taylors School in Ashwell but left at the end of 1910 having attained exemption age. He then went to work as a bottler at the local brewery. His parents had been proprietors of various pubs including the Fox in Ashwell, and the Saracen’s Head in Bedford.
Alfred did not marry and lived with his mother and sister in Bedford, Bedfordshire. He left half his estate to his sister Alice (Soames).
Alfred died at Shortstown, Bedfordshire on 5th March 1950.
Alfred’s brother Frank emigrated to Canada in 1910. It is not known if he served in the War.
Birth Place:
Ashwell, Hertfordshire
Enlistment Place:
Hertford, Hertfordshire
Royal Field Artillery
Battalion:
84th Army Brigade
Regimental Number:
Type of Casualty:
Theatre of War:
Western Europe, France and Flanders
Roll of Honour -WW1
Frederick Charles Walker
Edward Webb MM
Ernest Webb
Ernest William Westerman
Herbert George Westerman
Charles Hall Westrope
Ashwell Village Museum Heritage Fund
Major Fund Raising Initiative
Historic Diary - Ashwell Month by Month
Events from the pages of the Royston Crow
Ashwell Museum > http://www.ashwellmuseum.org.uk/
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Any of you read the Michael Moore letter to Bush?
Thread: Any of you read the Michael Moore letter to Bush?
abound media
anti-intro coalition founder
Phx AZ
I am curious to hear what many of you think about what he had to say in the letter. Regardless if you think he is a liar for some of his point in Bowling for Columbine, this letter seems to be right on the money. Atleast in my opinion??
Im anxious to hear what others think from both sides. Im quite open minded to new and differnt points of view.
Here is a link if you haven't seen or heard it on TV.
http://www.michaelmoore.com/words/index.php
japangreg
Didn't do it.
\o/ |o| |o_ /o\
``I don't remember seeing him. That does not mean he was not there,'' said Wayne Rambo, who was a first lieutenant with the 187th Supply Squadron at the time. The AP contacted more than a dozen former members of the unit on Wednesday, and none could recall ever running into Bush.
Anyone wanna argue with Rambo? Didn't think so.
Meanwhile, a retired Texas National Guard officer said Wednesday he overheard a conversation in 1997 between then-Gov. Bush's chief of staff, Joe Allbaugh, and then-Adjutant Gen. Daniel James of the Texas Air National Guard in which he contends those two men spoke about getting rid of any military records that would ``embarrass the governor.''
Retired Lt. Col. Bill Burkett told the AP that he saw documents from Bush's file discarded in a trash can a few days later at Camp Mabry in Austin, Texas. Burkett described them as performance and pay documents. He said the documents bore the header: ``Bush, George W. 1lt.'' -- meaning first lieutenant.
(both quotes from here)
Moore has moments of insight in his writing (as was shown in 'Dude, Where's my Country?') but I often find he crosses the line and starts to sound just plain whinny. He asks good questions, but the tone of his letter is a bit too confrontational for me to believe that he doesn't already have his own conclusions for this matter drawn up. In short, he isn't asking questions, he's making accusations with having to use a period.
That being said, I am not a Bush supporter; I dislike his policies, his mannerisms and his cocky sneer. I don't trust him at all, and I'll be voting against him in the elections.
I like the current pressure being put on Bush, not only from the AWOL issue but the discretionary spending issues, the pre-war intelligence failures, etc. What a change in the political climate from a year ago when criticizing the man was greeted like burning the flag while defiling mom's apple pie. It shows a recovering sense of accountability in this country that had all but vanished after 9/11.
Another good article on this issue: Yeoman of the Guard
Hush child. japangreg can do what he wants. - PAlexC
That was Zen - this is Tao.
Very well said, I can see where you're coming from. I had not read anything in regards to the NYtimes article. Atleast from the point of view that they share.
It's actually an AP piece, before someone starts bashing the Times as a liberal rag...
...would Jason Blair lie to you?
I'm the good one!
Ahh election year.....the mud slinging starts, the dirt is dug up-or made up-...I guess Bush will have to go a bit further in time than his opponent, possibly world war II since Kerry is about 63.
I mean talk about trivialities, did he serve in Alabama or not? man if thats the best they can do at this stage... Now who was it that dodged the draft---entirely??
Originally posted by XU1
Yes, election years are great; lace up those gloves, gonna be a bloody fight.
I assume you're talking about Clinton: I find this funny, seeing as it follows a comment about going back in time (Clinton isn't running again, is he? Then why bring it up?)
I think this is a very pertinent question, since Bush keeps wrapping himself in the mantle of 'war president' during interviews. How far can you really trust someone that tells you he is all about national defense when he skips out on 6-months of rather cushy duty when the nation was at war?
Now who was it that dodged the draft---entirely??
Dick Cheney?
//openedSource
Originally posted by japangreg
Because it shows that it didnt hurt Clinton at all...so the tactic is a no hoper...
also in this case the question is..."Where was he"?... the latest dental records show that he was there...I mean he could have been on light duties, or in the Kitchen...or sweeping the pavements...at least he enrolled..
Originally posted by CNO
I though it was Liza Minelli... ..but..Was this man ever president??
Not true when there is an ongoing war. Clinton wasn't hurt because the country was in a state of calm, enjoying a good economy.
Bush is in the White House during a time of domestic and international turmoil, with an open-ended, unconventional 'war' and nation building, and has sat though one of the worst economies in decades.
Clinton didn't have to have any war credentials; the country didn't need a military leader during his presidency.
If Bush is going to keep pushing the war as the main reason to keep him in office, he better be able to show that when given the light responsibility the ANG represented during Vietnam he didn't just act like a spoilt brat and drop the ball.
This AWOL issue is not a tactic to slander Bush as a whole. It is QUITE relivent today when he has sent our children and friends overseas. How many hundreds of people in the national gaurd have died in the last 9-11 months?
This is why it is important, what kind of leader is he if he cant set the example for the nation. If he doesnt have the balls to serve than why should anyone else? Its like telling your kid that swearing is bad and only to turn around spewing those same words from your mouth.
This is why it is important. In the Tim Russert interview this passed weekend, Bush acknowledged that the topic is brought up around any election he is in. However no one made it an issue before. Why you ask? BECAUSE WE WEREN'T AT WAR and at that point 600 of our kids were still breathing and enjoying there youths.
btw, it doesnt help that he has been unable to find WMD on land that we have control of. The only thing that any of us can truly proove about this war is that we captured Saddam (which I am thankfull for) and Cheney has become one hell of an oil Tycoon.
Last edited by abound media; 02-12-2004 at 05:37 PM.
TallGuyLittleCar
I Mastered Dead Technology
looking for my lighter
[B]Not true when there is an ongoing war. Clinton wasn't hurt because the country was in a state of calm, enjoying a good economy.
actually we were in the same war we are now in during the clinton administration. Clinton just choose not to fight, not suprising from a man who once said he despised the military.
can't help what you inherit. I suppose by that thinking abused children are responsible for the abuse they recieved.
So the first world trade center bombing was cool? the cole was cool? the embassy bombing in africa were fine. 9-11 was no big deal?
So you think Franklin Roosevelt was the wrong guy for ww2?
ONLY RON PAUL AND ALUMINUM FOIL CAN SAVE YOU NOW!
annoy your politician fairtax.org, a political forum
Catapultam habeo. Nisi pecuniam omnem mihi dabris, ad caput tuum saxum immane mittam.
Retired Mod
Originally posted by TallGuyLittleCar
just interested... how does that relate to the comment you quoted above it? Did I miss something?
C'mon now, all of you're responses here are aimed at bashing Clinton for not attacking Al Queda.
Neither did Bush. Not until we got hit.
And all the conservative sobbing over inheriting a bad economy doesn't excuse the problems that have occured on the Bush watch: even if we forgive him 9/11, the administration is directly responsible for the Iraqi war and the mishandling of the information. They are responsible for passing tax cuts while the nation's debt soars, underfunding their own educational programs, encouraging suspision of our intentions in Iraq by awarding no bid contracts... the list really does go on and on.
The point of this letter (remember the letter? The subject of this post? The one that doesn't mention Clinton at all?) was to ask a 'war time' president about his military service record, which appears to have several discrepancies.
You can try to blame Clinton for that all you like, but I believe he was in the UK at the time...
Originally posted by aversion
I think he's saying that we did need a war time pres during that time, to deter/prevent what happened.
Off the mark, TGLC?
said it better than i did, hence this edited post
Bush was handed a very difficult set of cards since 9/11...no one on earth could have envisioned the changing nature of terrorism, or how determined these people were/are to bring the fight to the US.
I really dont think that people will see Bush as some sort of Coward...
I just think Bush sees the world situation as it is.....I think he shows far sightedness....which means tough decisions, that are not going to be popular..
And if your going to bring the Hypocrisy issue up....does this mean that everytime a president needs to make a decision he has to stop going ahead with it because of some past mistakes or wrogdoing?
Its been a long time since he was in the service to be bringing this type of issue up...
once again at least he enrolled.
He could have been hiding behing some trees for six months..
Last edited by XU1; 02-12-2004 at 05:57 PM.
That was wierd; I went to quote something TGLC said ('...he's either doing a great job or a terrible job'... something like that), and when the form came up, it had his edited reply.
Spooked me.
I was going to say that that is true for just about everyone...
even if we forgive him 9/11,
???...Ok I give up now...I thought that some one else was responsible for 9/11...
[B]And if your going to bring the Hypocrisy issue up....doese this mean that everytime a president needs to make a decision he has to stop going ahead with it because of some past mistakes or wrogdoing?B]
Only when he makes a decision on that issue. Would you take hair care advice from a bald barber? How about being told to floss by a toothless dentist?
How does someone who ducked out of a cushy stateside post during a war where thousands of US service men are killed have the authority to ask other Guardsmen to risk and give their lives for something?
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Displaying items by tag: Blogonomics
What's New at BoomBustBlog
Those Who Are Not Familiar with Reggie Middleton's Work...
Can now learn about his many accurate and timely calls including every major market crash and nearly every major corporate collapse over the last 3 years. Also available, social media links and recent media appearances with streaming video. See More About Reggie Middleton.
How Has BoomBustBlog Research Done for the 3 Quarters of 2010?
Crains NY ran a happy, go lucky article today:
The stubbornly dismal economy means at least one thing: an extended stay in the spotlight for a handful of star analysts whose defining characteristic is their extraordinary bearishness. And, of course, their accuracy.
There's Albert Edwards, a London-based analyst from France's Société Générale, who believes the Standard & Poor's 500 will sink to 450, a sickening 57% drop from its current level. There's David Rosenberg, chief economist at Toronto money manager Gluskin Sheff, who warns that deflation is going to pull down the U.S. economy for years.
And then there's the New York star of this gloomy show: Reggie Middleton, a Brooklyn entrepreneur who turned to analyzing global markets after a stint buying and renovating apartments in Fort Greene and Clinton Hill. (See “Prophet of doom,” April 19.)
Bad as things may be for the economy, Mr. Middleton warns that they're poised to get much worse. Prices of real estate, stocks and bonds are all headed for serious falls... Wages will decrease, unemployment will increase. Fun, eh?
The culprit, Mr. Middleton says, is Washington. The bank bailouts, nationalization of Fannie Mae and Freddie Mac, and other interventions during two presidencies prevented the market from bottoming out in 2009 like it should have, he says. Now that the economy is weakening again and the heavily indebted U.S. government has fewer rescue options, the reckoning is coming. Markets of all kinds in the United States and Europe will get hit—hard.
“In my opinion, the amount of risk in the system is even higher than in 2008,” he says, adding this rare dash of hope: “2013 might be a good time to start taking a look at buying assets again.”
Mr. Middleton has been startlingly accurate in the past. He forecast the collapse of the housing market in 2007, and in early 2008 warned of the demise of Bear Stearns weeks before it happened. Earlier this year, he said that Ireland's finances were in terrible shape long before Standard & Poor's got around to downgrading that nation's credit rating.
For those of you who don't follow my blog, Mr. Elstein (the article's author) was referring to:
Does Pessimism Now Sell As Well As Sex, or Are Investors Just So Starved for the Unconflicted Truth?
Many have labeled me a Permabear, particularly my detractors and those in the media (who are decidedly not detractors but still paint a pessimistic bent on my outlooks, see sidebar below). I am nothing of the such. I am what will be soon be known as a realist, as opposed to being a pessimist or optimist. No, I am no Permabear. My proprietary investment style (see "The Great Global Macro Experiment, Revisited")
Crain’s New York
“His work is so detailed, so accurate, it’s among the best in the world,” says Eric Sprott, CEO of Sprott Asset Management, a Toronto firm that manages about $5 billion and subscribes to Mr. Middleton’s research.
Reggie in Forbes (Going short)
Middleton’s site combines self-promotion with meticulous financial analysis that is often delivered with a whiff of bathroom humor
dictates that I switch between extremes of bullishness and bearishness contingent upon the extreme policy errors of central bankers. As a matter of fact, I was as bullish as can be in residential real estate in NYC form 2000 to early 2004. I leveraged up on all the "in the money" distressed real estate I could find in areas of heavy gentrification, literally extracting 4 digit returns. That doesn't mean I disobeyed the laws of math though. As 2004 progressed, the writing on the wall became larger and more pronounced... Enter 2005 and math said turn bearish, common sense said turn bearish, and not to be one to look arithmetic in the face and argue, I grew bear claws and donned a ruffled brown grizzly coat! After liquidating my real estate, I took a year off and started shorting every industry that was even tangential to real assets. That was 2007. By the first quarter of 2009, I had a cumulative return in my portfolio of about 452% averaging roughly 50% cash (see Updated 2008 performance). I sensed the market was oversold, but the fundamental and macro outlooks was still quite negative, hence I pulled my profits one weekend in March (but let a few underwater positions ride). This weekend, coincidentally, happened to be the beginning of the rally that shouldn't have been, and I fought the faux bull to my detriment I got hurt in the artificially engineered, central banker and government synthesized rally of 2009, and my cumulative return was almost halved.
Who is Reggie Middleton and What is BoomBustBlog?
Reggie Middleton is an entrepreneurial investor who guides a small team of independent analysts to uncover truths, seldom if, ever published in the mainstream media or Wall Street analyst reports.
Reggie Middleton Wins CNBC's First Ever Stock Draft Investment Contest, and Does So By A Wide Margin!
Reggie Middleton's The Only Investor/Analyst To Publicly & Timely Call To Short Apple At It's All Time High and Go Long Google On CNBC!
The Financial Nostradamus!
Who Is Reggie Middleton & What Is BoomBustBlog?
Since the inception of his BoomBustBlog, he has established an outstanding track record, including but not limited to, the call of....
The housing market crash in the spring of 2006 and publicly in September of 2007: Correction, and further thoughts on the topic and How Far Will US Home Prices Drop?
The collapse of Bear Stearns in January 2008 (2 months before Bear Stearns fell, while trading in the $100s and still had buy ratings and investment grade AA or better from the ratings agencies): Is this the Breaking of the Bear? | After the collapse, a prudent bullish call as well... Joe Lewis on the Bear Stearns buyout Monday, March 17th, 2008: "The problem with the deal is that it is too low, and too favorable for Morgan. It is literally guaranteed to drive angst from the other side. Whenever you do a deal, you always make sure the other side gets to walk away with something. If you don’t you always risk the deal falling though unnecessarily. $2 is a slap in the face to employees who have lost a life savings and have the power to block the deal. At the very least, by the building at market price and get the company for free!" | BSC calls are almost free and the JP Morgan Deal is not signed in stone Monday, March 17th, 2008 | This is going to be an exciting, and scary morning Monday, March 17th, 2008 | As I anticipated, Bear Stearns is not a done deal Tuesday, March 18th, 2008 [Bear Stearns stock goes from $1 and change to $10, front month calls literally explode from pennies to several dollars]
The warning of Lehman Brothers before anyone had a clue!!! (February through May 2008): Is Lehman really a lemming in disguise? Thursday, February 21st, 2008 | Web chatter on Lehman Brothers Sunday, March 16th, 2008 (It would appear that Lehman’s hedges are paying off for them. The have the most CMBS and RMBS as a percent of tangible equity on the street following BSC. The question is, “Can they monetize those hedges?”. I’m curious to see how the options on Lehman will be priced tomorrow. I really don’t have enough. Goes to show you how stingy I am. I bought them before Lehman was on anybody’s radar and I was still to cheap to gorge. Now, all of the alarms have sounded and I’ll have to pay up to participate or go in short. There is too much attention focused on Lehman right now. ) | I just got this email on Lehman from my clearing desk Monday, March 17th, 2008 by Reggie Middleton | Lehman stock, rumors and anti-rumors that support the rumors Friday, March 28th, 2008 | It appears that I should have dug deeper into Lehman! May 2008
The fall of commercial real estate in general (September of 2007) and the collapse of General Growth Properties [nation's 2nd largest mall owner] in particular (November 2007):
Will the commercial real estate market fall? Of course it will.
Do you remember when I said Commercial Real Estate was sure to fall?
The Commercial Real Estate Crash Cometh, and I know who is leading the way!
Generally Negative Growth in General Growth Properties - GGP Part II
General Growth Properties & the Commercial Real Estate Crash, pt III - The Story Gets Worse
BoomBustBlog.com’s answer to GGP’s latest press release and Another GGP update coming… (among over 700 pages of analysis, review the January 2008 archives or search for “GGP” for more research).
The collapse of state and municipal finances, with California in particular (May 2008): Municipal bond market and the securitization crisis – part 2
The collapse of the regional banks (32 of them, actually) in May 2008: As I see it, these 32 banks and thrifts are in deep doo-doo! as well as the fall of Countrywide and Washington Mutual
The collapse of the monoline insurers, Ambac and MBIA in late 2007 & 2008: A Super Scary Halloween Tale of 104 Basis Points Pt I & II, by Reggie Middleton, Welcome to the World of Dr. FrankenFinance! and Ambac is Effectively Insolvent & Will See More than $8 Billion of Losses with Just a $2.26 Billion
The overvaluation of Goldman Sachs from June 2008 to present): “Can You Believe There Are Still Analysts Arguing How Undervalued Goldman Sachs Is? Those July 150 Puts Say Otherwise, Let’s Take a Look”, “When the Patina Fades… The Rise and Fall of Goldman Sachs???“andReggie Middleton vs Goldman Sachs, Round 2)
The ENTIRE Pan-European Sovereign Debt Crisis (potentially soon to be the Global Sovereign Debt Crisis) starting in January of 2009 and explicit detail as of January 2010: The Pan-European Sovereign Debt Crisis
Ireland austerity and the disguised sink hole of debt and non-performing assets that is the Irish banking system: I Suggest Those That Dislike Hearing “I Told You So” Divest from Western and Southern European Debt, It’ll Get Worse Before It Get’s Better!
The mobile computing paradigm shift, May 2010: More on the Creatively Destructive Pace of Technology Innovation and the Paradigm Shift known as the Mobile Computing Wars! »
Reggie Middleton Singularly Moves The Irish Banking System, Apparently Motivates Top Banking Regulator To Resign
We believe Reggie Middleton and his team at the BoomBust bests ALL of Wall Street's sell side research: Did Reggie Middleton, a Blogger at BoomBustBlog, Best Wall Streets Best of the Best?
There are several ways through which you can interact directly with Reggie:
Mr. Middleton Receives Positive Press Coverage, Worldwide!
Reggie Middleton Featured in Property EU, one of Europes leading real estate publicatios
Reggie Middleton on the Peter Schiff Show Discussing Facebook
Reggie Middleton on Max Keiser ZIRP and Treasury Ponzi Scheme
Reggie Middleton takes a Bite out of Apple on his trip back from Abu Dhabi & Dubai
Reggie Middleton on Goldman Sachs Business Model
On Greek Bailouts
On the Greek Bailouts...
http://media.cnbc.com/i/CNBC/components/Syndicated%20Video%20Player/videomodule.swf?id=3000073979&pcode=cnbcplayershare&play=&base=http://plus.cnbc.com/stickers/partners/cnbcplayershare/" >
Interesting documentary on the rating agencies' effect on the sovereign debt crisis in Europe, produced by VPRO Tegenlicht out of Amsterdam. Reggie Middleton appears in the following spots: 4:00, 22:30, 40:00...
Reggie Middleton Discussing the Rating Agencies effect on Sovereign Europe
Reggie Middleton currently leading the CNBC Stock Draft Pick contest
Reggie Middleton Explains the Travails of the F.I.R.E. Sector on CNBC
Reggie Middleton interviewed on Russion Television's Max Keiser discussing re-hypothecation thourgh MF Global at 12:32 in the video
Reggie Middleton interviewed on Russion Television's Capital Account on MF Global, Goldmand Sachs and Vampire Squids
Reggie Middleton interviewed on Russion Television's Capital Account concernign the European debt crisis and bank contation
Reggie Middleton on Mas Keiser discussing BNP Paribas bank runs and the potential collapse of French and European banks
Reggie Middleton on Mas Keiser discussing Goldman Sachs, currency debasement and ZIRP poisoning US banks
Reggie Middleton as the Keynote Speaker at the ING Real Estate Valuation Seminar in Amsterdam
Reggie Middleton on Bloomberg TV's Fast Forward
Reggie Middleton discusses the fall of commercial real estate in the US
Another stint on Max Keiser discussing topics such as Goldman’s Facebook offering that never was, what happens when its the banks that walk away from a home, phantom banking profits that never were, and more shenanigans that are the tour de force that is today’s banking system and economy. To skip directly to the Reggie Middleton interview, move to 11:55 in the video.
Reggie Middleton on BNR Dutch News Radio
A short interview clip on BNR, in Dutch. Click here.
Reggie Middleton on CNBC's Squawk on the Street - 10/19/2010
Mr. Middleton discusses JP Morgan, bank risk and technology and is the only pundit in the financial media that we know of that called Apple's margin compression issues and did so successfully just hours before they reported! Click here or click below to see the video.
Reggie Middleton with Max Keiser on the Keiser Report and RT Television - Discussing JP Morgan, Derivatives, Fraudclosure and the US Oligarchy
Go to 12:20 in the video to see the portion with Reggie Middleton
Bloomberg TV: "The risk/reward ratio in commercial real estate does not look good!"
BBC World News: "It wasn't just Lehman Brothers: Regulatory Capture is the Term du Jour!"
Amsterdam's VPRO Backlight and Reggie Middleton on brutal honesty, destructive derivatives and the "overbanked" status of many European sovereign nations
BBC World News Today and Reggie Middleton on the Obama Administration's attempt to reign in the US banking system
Reggie on CNN!
The bank stress tests were a sham! I know it, you know it! Everybody knew it but played along with the game anyway...
Reggie Middleton on the Young Turks: Another Economic Meltdown Coming???
Reggie Middleton in Forbes (Going short)
Middleton's site combines self-promotion with meticulous financial analysis that is often delivered with a whiff of bathroom humor
Las Vegas Business Press
NY Daily News - Giants claim defunct firm Lehman Brothers owes team $300M
MarketWatch (Herb Greenberg Blog)
Las Vegas Review Journal (Allegiant Air)
Las Vegas Review Journal (WYNN)
CNN.com (Blogger Bunch) May 7, 2009- Will Banks Fail?
CNN.com (Money & Mainstream) May 14, 2009 (see above)
BBC World News Today (multiple appearances, see above)
Older Press Coverage and Media Appearances (samples no longer available)
Real Estate Finance Today
Interactive Week
Computer Shopper
Hedge Funds Post: Best Blogs 2008
SoUp9lMCxRg
The Mobile Computing and Content Wars: Part 2, the Google Response to the Paradigm Shift
This is an excerpt from part two of a multi-part series on the companies vying for dominance during the 3rd major paradigm shift in personal and enterprise technology over the last 30 years. This one will be a biggie (not smalls) and promises to create an investment behemoth out of the winner and relegate the losers to relatively niche markets. This is saying a lot considering the size of the companies participating in the battle for the pole position. I created this series to provide a truly objective, truly informed, and truly analytical (from an empirical perspective) knowledge source on this very important intersection in personal computing and distributed media. This series will end with a full BoomBustBlog style forensic report on the company we feel has the most to gain from these wars from an investor's perspective. Those who are not familiar with my hard-edged, yet objective analytical work should reference past performance and media appearances for a quick background.
It is imperative that readers first review “There Is Another Paradigm Shift Coming in Technology and Media: Apple, Microsoft and Google Know its Winner Takes All” before moving on so as to get a thorough background as to what is at stake, who the players are, and what mobile technologies are being released into the consumer and enterprise realm. This is a lengthy, meaty, objective and information packed post that was initially intended to go out to subscribers only (click here to subscribe to our research services). I welcome you to compare it to the research you find available from technology, financial and strategic advisory firms, including and particularly Goldman Sachs (click here to see what I mean, then return to this point to continue reading) and let me know whose analysis is more accurate, in depth and thorough (not to mention less expensive).
Despite an increasing proportionate share of licensing and other revenues, advertising will apparently continue to drive Google’s revenues in the medium-to-long term. The Company’s dominant share in the search engine market is expected to be maintained, with other competitors failing to gain the operative, technological and strategic influence to deter its pace of ad revenue growth. Of late, the Company has been initiating to broaden its revenue base as is evident from some of the recent and planned launches of newer products and services funded through revenues generated by its online ad programs
Google’s next major launches – Chrome web browser (production version), Google Chrome OS and Google TV – (expected by the end of 2010) are expected by analysts to fare much better than its Nexus One phone. However, uncertainties over consumer acceptance, pricing and technological edge of newer services offerings over competing products in the market are making it quite difficult to gauge the near-term financial impact of these launches. I would like to make it known that it appears nearly all of the financial analysts and many of the technology analysts fail to grasp the gist of Google’s phone strategy. The Nexus One was a proof of concept, proven in the market place by HTC’s Evo, which (one month after being released) is constantly sold out in nearly all retail outlets, Sprint stores, indirect retails stores, and the Sprint.com site itself. It is also the first phone that is widely accepted as not only being a credible threat to the IPhone but actually superior to the IPhone as well. It should not be lost on anyone that the IPhone is what has driven the stellar burst in Apple’s revenues, profits and mindshare over the last three years. To that end, the Nexus One, and more aptly the open sourced Android OS has proven to be a raging success.
There Is Another Paradigm Shift Coming in Technology and Media: Apple, Microsoft and Google Know its Winner Takes All
With the cult-like adherence to Everything Apple, cultivated by the viral marketing engine that is Apple, it is very, very difficult to get objective comparisons and reviews of practically anything in a product category that Apple is present in. Yes, even the stalwart geeky tech site's have bitten the Apple, so to speak. Cnet, Wired, etc. are so Apple biased as to be borderline embarrassing. I know they report on what (and how whatever) brings the page views, but come on fellas!!! BoomBustblog thrives because it tells the truth in the financial and economic space, not matter how unpopular or controversial. Is it time to open up a BoomBustBlog, Technology edition????
What this abject bias does, despite irritating the hell out of those of us who are not plugged into the Matrix, is mask the exciting technical revolution that is occurring due to the intense competition borne from the weakening of the Wintel hegemony, the advent of a slew of new technologies across the telecomm, media, semiconductor and software industries and the new business models cropping up as the world finally embraces the World Wide Web as an actual permanent and primary platform for basic business, social and economic transactions.
This post will surmise the qualitative aspects of the companies and products listed below. I will follow up with full forensic analyses of not only the companies, but the business model and market share potential of each, as well as a thorough valuation scenario analysis. One of these companies will probably take over portable computing, and I think it will pay to hitch onto the right one. The next Microsoft is in the making. Hey, Microsoft may even be the next Microsoft. Don't sleep on them, although it does appear that they have been asleep themselves. We won't know until the Windows Mobile 7 OS is released. I recommend all who are interested in this tech, media or investments send the link to this article around the web, for it is one of the few (if not the only) truly unbiased reviews of the products that compete with Apple in the ultra portable and handheld space.
An Overview of the New Windows to the Web
From the top left hand corner, clockwise: The Amazon Kindle 2, Asus EEE PC, Apple IPad, Archos 5 Internet tablet, HTC Touch Pro2, Apple Itouch 3rd Generation, Archos 9 Internet Tablet, Sony Vaio. The paper under the Apple product is a testament to the viral marketing ability of Apple. My son did not want his ITouch to touch the floor!!!
A New Social Networking and Community System Has Been Installed
A new community and networking system has been installed. Please take the time to go through the features and take full advantage of it. You can upload and link your own videos, files, pictures charts, microblog, comment, network, etc. Each registered member an access it from their profile, or you can go to the community link at the top menu, also available here.
Hedge Funds Paying to Follow Bloggers: The Shift in Media Influence Continues
Here is an interesting article from thecrossbordergroup.com: Data firm monitoring influential blog sites
Hedge funds are testing out a data feed produced by monitoring a group of financial commentators that includes bloggers. The trial is still at an early stage, but it shows how seriously the investment community is taking the opinions of those working outside traditional media outlets. The feed – put together by Alacra, a data aggregator – monitors the output of around 25,000 analysts by combing the web for their comments. This group contains a number of prominent bloggers, such as Barry Ritholtz, Henry Blodget, Felix Salmon and Fred Wilson.
Are Blogs Truly Competitive With the Mainstream Media in Terms of Quality of Content?
A very interesting article was published in Crain's New York this morning, both in print and online, titled Prophet of doom. The article was about blogs, and the potential for them to raise capital and compete with the mainstream media. There's also a picture of a devilishly handsome, charismatic, and outright cute blogger there as well :mrgreen:
Here are a few excerpts, combined with my usual commentary...
Mr. Middleton's Boom Bust Blog forecast with stunning accuracy the demise of such real estate bubble blowups as Bear Stearns and mall operator General Growth Properties... Now he's embarking on his next project: Turning his blogging hobby into a full-fledged investment research business, a firm where he says investors will get “realistic” insights as opposed to Wall Street puffery. Mr. Middleton plans to start petitioning venture capital firms, private equity players and established media companies in the coming weeks.
Yes, it's official. I'm going to start building the blog out into something much bigger, more in depth, and more accessible.
“His work is so detailed, so accurate, it's among the best in the world,” says Eric Sprott, CEO of Sprott Asset Management, a Toronto firm that manages about $5 billion and subscribes to Mr. Middleton's research. Well, thank you Mr. Sprott! I owe you a good bottle of wine for that one! For those that don't follow Eric Sprott's commentary (or his investment record), this accomplished man tells it like it is. He was just on CNBC and rang the closing bell last week.
Finding investors won't be easy, though. A few bloggers have landed backers recently, including Footnoted.org, acquired by investment research firm Morningstar, and Gothamist.com, by Cablevision. But most media concerns and venture capitalists have steered clear. Many blogs' revenues are scant, with few opportunities to grow and churn out the $10 million to $15 million in revenues that venture capitalists want. “The climate for blogs is almost always lousy,” says Roger Ehrenberg, founder of IA Capital Partners, which has invested in BlogTalkRadio and BusinessInsider.com.
Do Blogs Compete at a Level that Threatens Mainstream Media?
I received an email (posted below) from Aaron Krowne, a fellow independent blogger and founder of the Implode-a-Meter group of web sites. His blog has broken many stories in the banking and hedge fund sector, and has a fairly broad network of independent sources. I am not commenting on the validity of the assertions and accusations herein, but I am casting my opinion on the apparent attempt to belittle the credibility of the blogs.
I can speak first hand on this topic. The specialized financial and political blogs offer expertise, data and analysis that often rival the best think tanks, banks and research houses in the world. I am not exaggerating here. I happen to be quite proud of the stuff that is generated. I am also impressed at the speed and actual accuracy of the breaking stories. This depth, collective breadth, and reflexive responsive times available through the blogoshpere makes it not only a viable conduit for news, analysis, opinion and information, but also an apparent threat to the struggling mainstream media model. Reference the back and forth between CNBC and Zerohedge as an example.
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