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(Continued) (A) Fas death domain structural model demonstrating six antiparallel α-helices (PDB: 1DDF) (19) with the side chains of Y232 and Y291 indicated (left) and amino acid sequences of human Fas and rat Fas encompassing the death domain with positions of each helix indicated (right; α, α-helix). Tyrosine phosph...
10.3389/fimmu.2016.00429
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FUnDinG This work was supported by institutional funds from the Centre National de la Recherche Scientifique (CNRS) and the Institut National de la Santé et de la Recherche Médicale (INSERM) , and by grants from the Institut National du Cancer (INCa ; PLBIO09-317 ); the University of Nice , the Agence Nationale ...
10.3389/fimmu.2016.00429
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FUnDinG This work was supported by institutional funds from the Centre National de la Recherche Scientifique (CNRS) and the Institut National de la Santé et de la Recherche Médicale (INSERM) , and by grants from the Institut National du Cancer (INCa ; PLBIO09-317 ); the University of Nice , the Agence Nationale ...
10.3389/fimmu.2016.00429
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Biochemistry, Genetics and Molecular Biology
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Introduction In the last 10 years, the prevalence of alternative splicing has been completely re-evaluated. Recent reports claim that more than 90% of multiexon genes produce at least two splicing variants [Pan et al. , 2008 , Wang et al. , 2008]. The depth at which we can sample transcriptomes with next generation se...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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, 2012 , Wang et al. , 2010 , Robertson et al. , 2010]. The ultimate goal of these methods is to identify and quantify full-length transcripts from short sequencing reads. This task is particularly challenging and recent benchmarks show that all methods still make a lot of mistakes [Steijger et al. , 2013]. The diffic...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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, 2014 , Freyermuth et al. , 2016]. The generalization of de novo assembly approaches for studying splicing in human seems to be mostly impeded by the lack of a clear evaluation of its potential in comparison to more traditional mapping-based approaches. This is the gap we aim at filling with the work presented here. T...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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The majority of frequent isoforms are found by both approaches Applying KisSplice and FaRLine to the same RNAseq dataset (SK-N-SH cell lines treated or not with retinoic acid) generated by the ENCODE consortium, we noticed that 68% of the alternatively skipped exons (ASE) identified by KisSplice were also identified by...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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Either the exon is unannotated (28%), one of its flanking exon is unannotated (8%) or both exons are annotated but no transcript combining them was annotated (45%). The assembly-first approach, KisSplice, does not consider annotations, and an interesting resulting advantage is that novel junctions have the same chance ...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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Overlap with other methods In a first step, we picked FaRLine and KisSplice as examples of a mappingfirst and an assembly-first approach respectively. Clearly, there are other published methods in both categories. MISO is probably the most widely used to annotate AS events. We therefore ran it on the same dataset to ch...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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Beside exon-centric approaches, which aim at finding the differentially spliced exons, there is also a number of published methods which are isoformcentric and have the more ambitious goal to reconstruct full-length tran-12 scripts. The most widely used mapping-first and isoform-centric approach is Cufflinks [Trapnell...
10.1101/074807
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2,016
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Biochemistry, Genetics and Molecular Biology
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KisSplice KisSplice is a local transcriptome assembler. As most short reads transcriptome assemblers [Grabherr et al. , 2011 , Schulz et al. , 2012 , Robertson et al. , 2010] , it relies on a De Bruijn graph (DBG). Its originality lies in the fact that it does not try to assemble full-length transcripts. Instead, it a...
10.1101/074807
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2,016
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Biochemistry, Genetics and Molecular Biology
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Annotation and quantification of alternative splicing events We wrote custom perl scripts, based on the FasterDB-based EnsEMBL r75 annotation database. For each gene, all the reads with at least one base overlapping the gene from the start to the end coordinates are retrieved. CIGAR strings are then used to retrieve th...
10.1101/074807
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2,016
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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This rule led to miss a lot of ASE events. So to define the flanking exons, we use the information contained in the transcripts and in the reads. We list each junction skipping an exon covered at least one read. If this junction is annotated in the transcripts, we extract all annotated events containing this junction. ...
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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SKNSH dataset We downloaded a total of 959M reads from http://genome. crg. es/encode_ RNA_dashboard/hg19/. They correspond to long polyA+ RNAs generated by the Gingeras lab, and are also accessible with the following accession numbers (ENCSR000CPN -SRA: SRR315315, SRR315316 and ENCSR000CTT -SRA : SRR534309, SRR534310)....
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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, FaRLine, use the FasterDB-based EnsEMBL r75 annotation database. FasterDB is a database containing all annotated human splicing variants [Mallinjoud et al. , 2014].
10.1101/074807
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Biochemistry, Genetics and Molecular Biology
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Acknowledgments This work was funded by the ANR-12-BS02-0008 (Colib'read) by the ABS4NGS ANR project ( ANR-11-BINF-0001-06 ), Action n3. 6 Plan Cancer 2009-2013, Fondation ARC ( Programme Labellisé Fondation ARC 2014 , PGA120140200853 ) and INCa ( 2014-154 ). Doctoral fellowships from ARC 1 -Région Rhône-...
10.1101/074807
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2,016
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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KRT8 was predominantly expressed in ovarian surface epithelia of controls (Figure 3I and 3J ). However, a focal distribution of KRT8 in TGFBR1-CA Acre ovaries was evident (Figure 3M and 3N ). Double immunofluorescence revealed low expression or absence of INHA in KRT8-positive cells within tumor lobules (Figure 3O...
10.18632/oncotarget.10149
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Biochemistry, Genetics and Molecular Biology
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Hormone profile of mice harboring a constitutively active TGFBR1 in the ovary Clinically, granulosa cell tumors often secrete excessive inhibin and estrogen [32, 33]. Serum inhibin is a reliable marker for granulosa cell tumors [34]. To determine the relevance of TGFBR1-CA Acre mice as a potential model for granulosa...
10.18632/oncotarget.10149
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Biochemistry, Genetics and Molecular Biology
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Histological analysis Ovarian samples were collected from control and experimental mice and processed for histological analysis. A standard protocol was used for hematoxylin and eosin (H&E) staining. Microscopic images were captured using DP25 (Olympus) digital camera interfaced with cellSens Imaging Software.
10.18632/oncotarget.10149
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Biochemistry, Genetics and Molecular Biology
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Immunofluorescence microscopy and immunohistochemistry Immunofluorescence and immunohistochemistry were performed using serial paraffin sections (5 μm) [18] , with primary antibodies listed in Table S1. Secondary antibodies for immunofluorescence were conjugated with Alexa Fluor 488 or 594. The sections were mounted ...
10.18632/oncotarget.10149
article
en
2,016
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Biochemistry, Genetics and Molecular Biology
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Statistical analyses All statistical analyses were performed using IBM Statistical Package for the Social Sciences (SPSS; Version 23) except the survival rate analysis by Mantel-Cox test using GraphPad Prism 6 software. Two sample independent t-test (unpaired) was performed to determine difference between two means. Th...
10.18632/oncotarget.10149
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Biochemistry, Genetics and Molecular Biology
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ACKNOWLEDGMENTS We thank Drs. Robert Burghardt and David Threadgill for insightful comments and critical reading of the manuscript. We thank Dr. Aline Rodrigues Hoffmann for helpful comments and discussions. We thank Drs. JoAnne Richards and Jan Gossen for the Cyp19-Cre mice and Dr. Richard Behringer for th...
10.18632/oncotarget.10149
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2,016
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Biochemistry, Genetics and Molecular Biology
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, 2001). PCR products were visualized after electrophoresis using a 2% agarose gel, staining with ethidium bromide (at 5 mg/L for 15 min) and UV light exposure. PCR products were sequenced by the Biofidal (Villeurbanne, France) sequencing platform. Their DNA sequences were analyzed using the BioEdit software. Similarit...
10.3389/fmicb.2016.00811
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2,016
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Biochemistry, Genetics and Molecular Biology
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(1996). Neighbor-joining trees and 1000 bootstrap replicates were generated. The adjusted Wallace coefficients enabling comparisons of partitions between genetic markers (lec and MLST types) were computed according to Severiano et al. (2011) using UMMI web site at Universidade de Lisboa (http://www. comparingpartitio...
10.3389/fmicb.2016.00811
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2,016
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Biochemistry, Genetics and Molecular Biology
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Comparative Genomics Assembled genomes of three PA7-related strains, i. e. , EML528, EML545 and EML548 used in this study have been deposited at DDBJ/EMBL/GenBank under the following accession numbers: LFXS00000000. 1 for EML528, LGJE00000000. 1 for EML545, and LFXR00000000. 1 for EML548 (Boukerb et al. , 2015). These...
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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Microcalorimetry Recombinant LecB PA7 was dissolved in a 20 mM TrisHCl, pH 7. 5, NaCl 150 mM, 100 µM CaCl 2 buffer. Carbohydrate ligands were dissolved in the same buffer, degassed, and loaded in the injection syringe. ITC was performed using ITC200 microcalorimeter (Malvern Instruments, UK). The LecB PA7 solution was ...
10.3389/fmicb.2016.00811
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2,016
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Biochemistry, Genetics and Molecular Biology
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Protein Crystallography Crystals of LecB PA7 complexed with Lewis a tetrasaccharide (Elicityl, Crolles, France) were obtained by the hanging drop vapor diffusion method using 2 µL of drops containing a 50:50 (v/v) mix of protein and reservoir solution at 19 • C. The protein at 10 mg/mL in 20 mM HEPES pH 7. 5 and 1 µM C...
10.3389/fmicb.2016.00811
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2,016
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Biochemistry, Genetics and Molecular Biology
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Water molecules, introduced automatically using Coot, were inspected manually. The stereochemical quality of the models was assessed with the program Molprobity (Chen et al. , 2010) , and coordinates were deposited in the Protein Data Bank under code 4UT5.
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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Bacterial Aggregation Assays through Imaging Flow Cytometry Analysis Aggregation assays were performed according to Boukerb et al. (2014) but aggregates were visualized and analyzed by Amnis FlowSight R cytometry. Image acquisitions and analyses were performed with the ImageStream system (Amnis Corporation, Seattle, ...
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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RESULTS Prevalence and Diversity Of lecA and lecB lecA and lecB databases were built by retrieving the P. aeruginosa sequences available at GenBank and at the Pseudomonas Genome database (Winsor et al. , 2015) and by performing lecA and lecB PCR screenings over a collection of environmental and clinical strains (Figu...
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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Comparison of Affinities between LecB PA7 and LecB PAO1 Purified LecB PA7 produced in E. coli were assayed on the Glycan Array v5. 2 from the Consortium for Functional Glycomics (n = 609 glycans). Only glycans with terminal fucose or mannose residues were recognized by the lectin. Stronger signals were obtained for bia...
10.3389/fmicb.2016.00811
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false
Biochemistry, Genetics and Molecular Biology
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Some of these RGP matched DNA segments smaller than 5 kb but were confirmed, in some instances, to be targets for longer genetic elements e. g. , RGP90. The lecB loci genomic environments were not subjected to similar large scale rearrangements but rather to small ones (up to 16 kb). The α genomic region harboring lecA...
10.3389/fmicb.2016.00811
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true
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false
Biochemistry, Genetics and Molecular Biology
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LecB cluster III strains yielded significantly higher numbers of aggregates when exposed to Calix-Fuc 4 and Calix-Glc 4 than monovalent sugars (Figure 7 and Supplementary Figure S12 ). However, significant differences could not be obtained using Calix-Man 4 even though, in average, higher numbers were measured. Inte...
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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These assays were performed on P. aeruginosa strains of (A) LecB cluster I (PAO1, its isogenic mutants PAO1 lecA and PAO1 lecB, bpoe1643 and GR23), (B) cluster II (PA14, bpoe1392, bpoe1737 and bpoe1748), and (C) cluster III (PA7, EML528, EML545 and EML548) (according to Figure 1 ). Aggregation levels were recorded fro...
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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ACKNOWLEDGMENTS We thank Sophie de Bentzmann and David Redelberger of UMR CNRS 7255 , Aix Marseille Université , for having made available the PAO1 lecA and PAO1 lecB mutants. The glycan array resources were provided by the Consortium for Functional Glycomics ( GM62116 ).
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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FUNDING This work was partly funded by Anses project " pyo-eau " # 2011/1/137 of the " Programme Environnement-Santé-Travail " (French Ministers in charge of ecological and environmental issues). We thank the CNRS, Université Lyon 1, VetAgro Sup, Cluster "Infectiology" of the Rhône-Alpes region, and Labex IMU (Inte...
10.3389/fmicb.2016.00811
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Biochemistry, Genetics and Molecular Biology
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The tip Triple Mutant Exhibits Shorter Roots at 11 d after Germination Since AtTIP genes are expressed in elongation zones, the lack of the most highly expressed isoforms could result in a deficiency in cell elongation. As we did not observe any difference in hypocotyl elongation between the different tip mutant lines ...
10.1104/pp.15.01635
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Agricultural and Biological Sciences
https://openalex.org/fields/11
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In some seedlings of the tip1;2 tip2;1 and tip1;1 tip1;2 tip2;1 mutants, LRPs still remained in stage II, in contrast to the wild-type plants, where the earliest LRP stage found was V. These data strongly indicate an important role of AtTIPs in LRP development. The nonadditive response of the single, double, and triple...
10.1104/pp.15.01635
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Agricultural and Biological Sciences
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Plant Material and Growth Conditions The Arabidopsis (Arabidopsis thaliana) tip1;1 mutant is a T-DNA insertion mutant from the Salk Institute collection (Alonso et al. , 2003) and was obtained from the Nottingham Arabidopsis Stock Centre (NASC) (seed stock N519003). The Arabidopsis tip1;2 and tip2;1 mutants were prev...
10.1104/pp.15.01635
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Agricultural and Biological Sciences
https://openalex.org/fields/11
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Accession Numbers Sequence data from this article can be found in the Arabidopsis Genome Initiative of GenBank/EMBL databases under the following accession numbers: AtTIP1;1, At2g36830; AtTIP1;2, At3g26520; AtTIP1;3, At4g01470; AtTIP2;1, At3g16240; AtTIP2;2, At4g17340; AtTIP2;3, At5g47450; AtTIP3;1, At1g73190; AtTIP3;2...
10.1104/pp.15.01635
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Agricultural and Biological Sciences
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ACKNOWLEDGMENTS We thank the imaging platform IMABIOL. Received October 20, 2015; accepted January 19, 2016; published January 22, 2016.
10.1104/pp.15.01635
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Agricultural and Biological Sciences
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V iewers "know" how to visualize their target classes and will produce a list of V iews. Each V iew can then be displayed in one or more environment, such as the desktop or web application. opens a project and select a given protocol, fills its parameters and execute it. (B) Inputs selected by the user are validated an...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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The introduction of direct detection devices (DDDs) has made a fundamental difference in image acquisition quality, enhancing the resolution achieved by previous image-recording media, such as photographic films or charge-coupled devices. Moreover, computer power has increased significantly through the use of multi-cor...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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In this thesis I address essential software problems in the field including: interoperability, traceability, reproducibility, data management, automation and distributed computing. This work describes the design and implementation of a new software framework, Scipion, that models image processing in electron microscopy...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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One important aspect in the FSC definition is the assumption of the noise independence in the two maps. This condition is difficult to meet in practice and it is often compromised by refining a single dataset while evaluating the FSC with two volumes computed from half-subsets of the dataset. In 3DEM, the "resolution" ...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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Ease of Use and Extensibility The proposed solution should also benefit software developers in the 3DEM community by providing an integrative framework with built-in tools. The system should be easy to extend to allow quickly incorporation of new algorithms emerging in the field. In that sense, the task of adding a new...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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The SPIDER scripting mechanism allowed the development of applications to deal with a large number of images, which also involved a complex set of mathematical operations. Moreover, users could design the processing path through the use of batch scripts, even without a strong background in scientific programming. In SP...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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Other operations on document files, such as sorting, are also available. Originally SPIDER contained all the code for the system, including calls to various graphics devices. With the arrival of graphical user interfaces (GUI) and standardization on the Motif/XWindow system, it became difficult to integrate interactive...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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SPIRE monitors executing processes and, after finishing, checks if expected outputs where generated and add them to an internal project database. The list of successfully executed batch files is displayed in the Project Viewer, as well as output files generated by each one. When a new project is started in SPIRE, a dia...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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State of the Art but only if the script contains a header following certain rules. Apart from the benefit of the generated GUI, the header constrain encourage the good practice of separating input/output variables from the processing commands in the batch file. SPIRE stores the output files into an internal project dat...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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It is also possible to generate HTML files from a project, which is specially useful at the end of a project as a review of the processing history. Another interesting feature is the possibility to import data from an existing project database, as well as export when the project is finished.
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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It also supported parallel processing on clusters, SMP supercomputers or sets of individual workstations. Despite the package had a tiered architecture, after many years of development (original in Objective-C, then C++ and later Python bindings) there was no clear organization to incorporate new features [47]. The ne...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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Another huge re-factoring of the package was finished in 2013, which led to a new major release, Xmipp 3. 0 [54]. This version introduced notable improvements in terms of project management, core libraries, GUIs, image formats and standardization. In Xmipp 2. 4 there was no clear concept of a project. While there was ...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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Previous to Xmipp 3. 0, the processing was protocol-oriented: the user launched scripts to perform the required operations. Communication between different protocols was achieved manually, which required that the user know what output files were needed as input for the next protocol. There was also no way to track work...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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Appion Till the introduction of Scipion, Appion was the only platform that allows real integration among software packages [45]. Appion is tightly connected with a previous project by the same group, Leginon [92] , a system for automated data acquisition that tracks and records all microscope parameters associated wi...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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One of the reasons to choose Python, a powerful and easy to learn language, was to quickly incorporate new tools into the pipeline. One of the cornerstones of the Appion system was the underlying database, which contained all the processing information and provided the links between disparate packages. The creation and...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
https://openalex.org/fields/13
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The Sinedon library was based on the concept of object-relational mapping (ORM), which refers to transferring objects from a programming language to a database system. With this library, new tables could be created on-the-fly from Python scripts, as well as launching different queries. Through Sinedon, Appion developer...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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py to store and retrieve data from the database. The general module appion/Loop. py iterates over a set of images in an experiment and pause the processing until a new image is entered in the database. This functionality allows to perform some operations (e. g. Particle Picking or CTF estimation) concurrently with the ...
10.1016/j.jsb.2016.04.010
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Biochemistry, Genetics and Molecular Biology
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Current research in EM depends on the analysis of huge amounts of data. These are in several formats and use different vocabularies. Furthermore, data are often stored or distributed using formats that leave implicit many important features relating to the structure and semantics of the data. It is really surprinsing t...
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In this case, a relational database is implemented for data processing but the data model is not clearly described. Another example is EMEN2 [95] , developed by the same group of EMAN2 [47] , that implements an electronic notebook using an object oriented database. This database is built around two concepts: protocol...
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Their authors claim that it is possible to incorporate portions of the processing pipeline, but currently tasks such as 3D reconstruction and refinement are treated as external processes. It seems that the ontology developed for EMEN2 is not directly used by EMAN2 for the implementation of image processing programs. Fi...
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Scientific Workflows Scientific workflows have been increasingly used in the last decade for data intensive science [97]. A variety of workflow systems, both open source (e. g. Taverna [98] , Galaxy [99] , Vistrails [100] , Kepler [101] , Triana [102] , ASKALON [103] ) and commercial (e. g. Pipeline Pilot2 [104...
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Most Taverna workflows are composed from a mixture of distributed Web Services, local scripts and other service types. Nevertheless, in some cases the installation only uses local services, where data and software are placed together and network traffic is notably reduced. One example of such scenario is the usage of T...
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Nonetheless, there is a large redundancy of web service functions that allows to identify reliable services and alternatives to make workflows more robust. The workflow engine in Taverna is implemented in Java as a multi-threaded object model, where processors are represented by objects, and data transfers are realized...
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Galaxy In 2005, the Galaxy Project started with the main goal of enabling researchers without programming and system administration expertise to perform computational analysis through the Web. It was designed as a system for the integration of genomic sequences, their alignments and functional annotations. Despite bein...
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The main window (shown in Figure 3. 3) is divided into three panels: the left one shows available tools grouped into categories that can also be searched; the middle panel is used to fill the parameters of the selected tool or to inspect an output dataset; the right panel contains the full history of actions and produ...
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Workflows are critical to automate and repeat the analysis of large datasets. Workflows capabilities have been improved since the first versions of Galaxy. For example, workflows can now be paused and restarted. Moreover, entire workflows can be embedded into another workflow as a single step, what facilitates the crea...
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The vistrails are stored using XML, which is a very flexible format to accommodate variability in the information being stored. A clear benefit of using an open and self-describing format is the ability to query, share and publish vistrails among scientists. The execution of a vistrail can take a long time depending on...
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This model is a cornerstone of the entire Scipion framework to integrate of existing EM programs and to adapt to future methodologies and requirements. Moreover, having a good specification of the processed objects allows to go one level up and concatenate more efficiently different steps to form an entire workflow. By...
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Design Evolution The initial Scipion design made use of a relational database (PostgreSQL) to represent the EM data model. Very soon we recognized that it was not a good approach due to the large number of tables required that were very difficult to maintain. Relational databases are very useful when the data model is ...
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This new approach was much more flexible to adapt to new data entities and algorithms. The new design was implemented using the Jena-TDB ontological database (http: //incubator. apache. org/jena/documentation/tdb/index. html) for metadata storage. The persistence layer used Empire (an implementation of Java Persistence...
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First, it was not easy to extend with new protocols since the classes (Java or Python) were automatically generated from the ontology definition. So, adding new protocols or data objects required an entire re-build of the ontology and code generation. This was not an easy procedure to be done each time a new protocol w...
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Movies The Movie class is a specialization of Image that contains the location of the movies frames stored in the hard disk and can store information related with frames alignment. By storing only the relative shifts between frames to align them, we can later obtain the aligned binary movie on demand. This allow us to ...
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Process Model Another cornerstone of our entire business model is the process model, which is complementary to the previously discussed data model. The central element of the process model in Scipion is the Protocol class, that can be defined as a container of algorithms and the engine for its execution. Protocol objec...
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Both patterns address the problem of dealing with an external system that is not under our control that we want to simplify or adapt to fit our desired interface. In our specific case, the Protocol class acts as a wrapper over the underlying programs. On one side, each protocol should be consistent with the Scipion int...
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If we want to apply the same processing strategy to a new dataset, we can simply export the workflow template from the previous project and import it into the new one. Once imported, the new runs will appear as new copies with the proper parameters and connections. We can execute them as usual or modify some steps of t...
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Streaming Processing The first Scipion release was designed so that if a run r2 depends on an object created by a run r1, then r2 could not be launched until r1 finished. Soon we discovered that, under certain circumstances, this approach introduced a performance limitation. For example, it prevented to execute pre-pro...
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For example, in streaming mode, the import protocol keeps monitoring the data folder to add more elements as they are acquired by the microscope. To support the data stream processing, some protocols were modified to adopt a more proactive behaviour to discover new input data, process it and update their output accordi...
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Packages As mentioned in previous chapters, Scipion was conceived with extensibility in mind. For EM we developed a basic data and process model to represent the processing pipeline, but Scipion design allows the extension of this model. In order to extend the current EM model, it is required to provide new packages (r...
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Protocols Together with the EM data model discussed previously, we also developed several EM protocol classes that serve as the base for further extension. The classes hierarchy diagram shown in Figure 4. 1 represents the main categories of protocols for EM. Protocols derived from P rotImport serve to incorporate exte...
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Viewers Another type of objects that could be defined in the framework is the V iewer, which handle the visualization of other objects such as protocols or data. Viewers are also important objects since visualization and data analysis are essentials for EM processing. In Scipion, a viewer class can be linked to one or ...
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Implementation This section describes the implementation of three important parts of the Scipion framework: (a) the persistence mechanism, (b) the graphical interfaces and, (c) the execution engine. Most Scipion code is written in Python since it is a very versatile scripting language. Python has been widely adopted fo...
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Automatic Data Persistence In 3DEM, most software suites use text files for metadata storage. A less popular alternative is databases. Leginon and Appion use the Sinedon library to map Python classes onto a relational database. The underlying database was implemented with MySQL (https://www. mysql. com/), which require...
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In Xmipp 3, the SQLite library was used internally to operate on image metadata. In Scipion, one of the design principles was to separate the conceptual model from the physical model. For achieving that goal, we looked into the object-relational mapping (ORM) concept, which allows data transfer between objects in a pro...
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Another pattern is the Data M apper [116] , in which there is a clear separation between the domain and the database and the objects do not have persistence methods. Both patterns have their pros and cons, as well as using ORM tools. One of the concerns that is associated with the use of ORMs is the potential for redu...
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Despite we have used Sqlite as the engine to store and retrieve objects, our implementation allows to replace this mapper with other approaches, either based on another database engine or even using text files (e. g, xml or json). In Scipion, we found two different contexts with different requirements for the ORM layer...
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SqliteMapper SqliteM apper was the first implemented mapper for storing/retrieving objects derived from Object (the root class in our Python basic hierarchy). One of the main requirements for this mapper is flexibility, because SqliteM apper needs to deal with objects that have a variable number of attributes. For exam...
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The set S 2 can be aligned and classified in 2D, from which one could create a subset (that also preserves the RELAT ION T RAN SF ORM ) S 3. Since we have the transformation path S 1 → S 2 → S 3 , we could extract some properties from The resulting SqliteM apper class fulfilled our requirement of flexibility, since it ...
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SqliteFlatMapper Since we work with many sets in a given EM project (micrographs, CTFs, particles, etc), and they could contains large number of items, we store each set separately of the project database. In this way we could load items on demand and reduce the risk of performance issues if the database grows too much...
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At this time, the mapper also computes the IN SERT and U P DAT E SQL statements that will be used for inserting and updating items. For each new object, its attributes are retrieved and stored into the database. This approach requires that all items stored in the set should be of the same class and contain the same att...
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In this table the attribute name and class are also stored to allow re-constructing the object from their recorded values in a given row. The Objects table has one row per item in the set, containing the values for the object attributes. The Figure 4. 4a shows the Classes table of a SetOf P articles database where the...
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Web Interface and Online Tools Online web applications and tools are common in Bioinformatics, providing easy access to algorithms without the need of local software installation. This is not the case for EM, mainly because most of the operations involve manipulation of huge datasets that are not easy to transfer. Neve...
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Execution engine The whole Scipion framework heavily relies on the underlying execution engine, i. e. , how jobs are setup and launched, or how protocol steps are executed. This section describes the mechanisms used by Scipion to facilitate jobs management for users, as well as for developer during protocols developmen...
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As mentioned in previous sections, if the input validation fails for a protocol, it is not launched and the errors are shown to the user (Figure 4. 11 2). If the validation is passed, Scipion prepares the conditions for the protocol execution. First, the project database is updated with the new state of this protocol ...
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oracle. com/us/products/tools/ oracle-grid-engine-075549. html). Each protocol is executed in an independent process, spawned either by the jobs management system or directly by the P roject process (Figure 4. 11 3). The new process will load input values and the project environment from its own database. Depending on...
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For each run, Scipion uses the steps. sqlite database to record the steps, their parameters and their status. When a protocol runs in Resume mode, the current list of steps is compared with the previous list (if exists) stored in steps. sqlite to determine which steps need to be executed. Scipion provides this re-start...
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In that case, Scipion can use either POSIX threads or MPI processes to execute steps concurrently. When using threads, the T hreadStepExecutor class is instantiated and it will spawn N threads (N is the number of parallel processes selected by the user) to work on the steps until the whole protocol is finished. There i...
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Optimization of CNB Microscope Acquisition Parameters After the Talos-Artica microscope was installed in the CNB, Dr. Daniel Luque used Scipion to run several experiments to calibrate some of the acquisition parameters and to find optimal parameters for the first steps of the workflow. As a test case, he used a Circovi...
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The obtained resolutions were similar at all frequencies, except for higher ones, where the 73k dataset seems to have a better resolution. Moreover, the pixel size was calibrated for the 92K dataset. It was found that from the nominal pixel size of 1. 14 A/px, the calibrated one is 1. 09 A/px. 2. Comparison of the sele...
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Comparison of different combination of programs for movie alignment: The same dataset of 58k particles (with 73K magnification) was used to compare the following combinations for movie alignment: (a) correlation plus optical flow, (b) motioncor2 and, (c) motioncor2 with dose compensation. For this comparison, all frame...
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