A Resource Optimized GATK 4 Based Open Source Variant Calling Workflow
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Sequence Alignment/Map) Is a Text Format for Storing Sequence Alignment Data in a Series of Tab Delimited ASCII Columns
NGS FILE FORMATS SEQUENCE FILE FORMATS FASTA FORMAT FASTA Single sequence example: >HWI-ST398_0092:1:1:5372:2486#0/1 TTTTTCGTTCTTTTCATGTACCGCTTTTTGTTCGGTTAGATCGGAAGAGCGGTTCAGCAGGAATGCCGAGACCGAT ACGTAGCAGCAGCATCAGTACGACTACGACGACTAGCACATGCGACGATCGATGCTAGCTGACTATCGATG Multiple sequence example: >Sequence Name 1 TTTTTCGTTCTTTTCATGTACCGCTTTTTGTTCGGTTAGATCGGAAGAGCGGTTCAGCAGGAATGCCGAGACCGAT ACGTAGCAGCAGCATCAGTACGACTACGACGACTAGCACATGCGACGATCGATGCTAGCTGACTATCGATG >Sequence Name 2 ACGTAGACACGACTAGCATCAGCTACGCATCGATCAGCATCGACTAGCATCACACATCGATCAGCATCACGACTAGCAT AGCATCGACTACACTACGACTACGATCCACGTACGACTAGCATGCTAGCGCTAGCTAGCTAGCTAGTCGATCGATGAGT AGCTAGCTAGCTAGC >Sequence Name 3 ACTCAGCATGCATCAGCATCGACTACGACTACGACATCGACTAGCATCAGCAT SEQUENCE FILE FORMATS FASTQ FORMAT FASTQ Text based format for storing sequence data and corresponding quality scores for each base. To enable a one-one correspondence between the base sequence and the quality score the score is stored as a single one letter/number code using an offset of the standard ASCII code. Quality scores range from 0 to 40 and represent a log10 score for the probability of being wrong. E.g. score of 30 => 1:1000 chance of error SEQUENCE FILE FORMATS FASTQ FORMAT FASTQ Each fastq file contain multiple entries and each entry consists of 4 lines: 1. header line beginning with “@“ and sequence name 2. sequence line 3. header line beginning with “+” which can have the name but rarely does 4. quality score line SEQUENCE FILE FORMATS FASTQ FORMAT FASTQ @HWI-ST398_0092:6:73:5372:2486#0/1 TTTTTCGTTCTTTTCATGTACCGCTTTTTGTTCGGTTAGATCGGAAGAGCGGTTCAGCAGGAATGCCGAGACCGAT -
The Variant Call Format Specification Vcfv4.3 and Bcfv2.2
The Variant Call Format Specification VCFv4.3 and BCFv2.2 27 Jul 2021 The master version of this document can be found at https://github.com/samtools/hts-specs. This printing is version 1715701 from that repository, last modified on the date shown above. 1 Contents 1 The VCF specification 4 1.1 An example . .4 1.2 Character encoding, non-printable characters and characters with special meaning . .4 1.3 Data types . .4 1.4 Meta-information lines . .5 1.4.1 File format . .5 1.4.2 Information field format . .5 1.4.3 Filter field format . .5 1.4.4 Individual format field format . .6 1.4.5 Alternative allele field format . .6 1.4.6 Assembly field format . .6 1.4.7 Contig field format . .6 1.4.8 Sample field format . .7 1.4.9 Pedigree field format . .7 1.5 Header line syntax . .7 1.6 Data lines . .7 1.6.1 Fixed fields . .7 1.6.2 Genotype fields . .9 2 Understanding the VCF format and the haplotype representation 11 2.1 VCF tag naming conventions . 12 3 INFO keys used for structural variants 12 4 FORMAT keys used for structural variants 13 5 Representing variation in VCF records 13 5.1 Creating VCF entries for SNPs and small indels . 13 5.1.1 Example 1 . 13 5.1.2 Example 2 . 14 5.1.3 Example 3 . 14 5.2 Decoding VCF entries for SNPs and small indels . 14 5.2.1 SNP VCF record . 14 5.2.2 Insertion VCF record . -
A Semantic Standard for Describing the Location of Nucleotide and Protein Feature Annotation Jerven T
Bolleman et al. Journal of Biomedical Semantics (2016) 7:39 DOI 10.1186/s13326-016-0067-z RESEARCH Open Access FALDO: a semantic standard for describing the location of nucleotide and protein feature annotation Jerven T. Bolleman1*, Christopher J. Mungall2, Francesco Strozzi3, Joachim Baran4, Michel Dumontier5, Raoul J. P. Bonnal6, Robert Buels7, Robert Hoehndorf8, Takatomo Fujisawa9, Toshiaki Katayama10 and Peter J. A. Cock11 Abstract Background: Nucleotide and protein sequence feature annotations are essential to understand biology on the genomic, transcriptomic, and proteomic level. Using Semantic Web technologies to query biological annotations, there was no standard that described this potentially complex location information as subject-predicate-object triples. Description: We have developed an ontology, the Feature Annotation Location Description Ontology (FALDO), to describe the positions of annotated features on linear and circular sequences. FALDO can be used to describe nucleotide features in sequence records, protein annotations, and glycan binding sites, among other features in coordinate systems of the aforementioned “omics” areas. Using the same data format to represent sequence positions that are independent of file formats allows us to integrate sequence data from multiple sources and data types. The genome browser JBrowse is used to demonstrate accessing multiple SPARQL endpoints to display genomic feature annotations, as well as protein annotations from UniProt mapped to genomic locations. Conclusions: Our ontology allows -
A Combined RAD-Seq and WGS Approach Reveals the Genomic
www.nature.com/scientificreports OPEN A combined RAD‑Seq and WGS approach reveals the genomic basis of yellow color variation in bumble bee Bombus terrestris Sarthok Rasique Rahman1,2, Jonathan Cnaani3, Lisa N. Kinch4, Nick V. Grishin4 & Heather M. Hines1,5* Bumble bees exhibit exceptional diversity in their segmental body coloration largely as a result of mimicry. In this study we sought to discover genes involved in this variation through studying a lab‑generated mutant in bumble bee Bombus terrestris, in which the typical black coloration of the pleuron, scutellum, and frst metasomal tergite is replaced by yellow, a color variant also found in sister lineages to B. terrestris. Utilizing a combination of RAD‑Seq and whole‑genome re‑sequencing, we localized the color‑generating variant to a single SNP in the protein‑coding sequence of transcription factor cut. This mutation generates an amino acid change that modifes the conformation of a coiled‑coil structure outside DNA‑binding domains. We found that all sequenced Hymenoptera, including sister lineages, possess the non‑mutant allele, indicating diferent mechanisms are involved in the same color transition in nature. Cut is important for multiple facets of development, yet this mutation generated no noticeable external phenotypic efects outside of setal characteristics. Reproductive capacity was reduced, however, as queens were less likely to mate and produce female ofspring, exhibiting behavior similar to that of workers. Our research implicates a novel developmental player in pigmentation, and potentially caste, thus contributing to a better understanding of the evolution of diversity in both of these processes. Understanding the genetic architecture underlying phenotypic diversifcation has been a long-standing goal of evolutionary biology. -
VCF/Plotein: a Web Application to Facilitate the Clinical Interpretation Of
bioRxiv preprint doi: https://doi.org/10.1101/466490; this version posted November 14, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Manuscript (All Manuscript Text Pages, including Title Page, References and Figure Legends) VCF/Plotein: A web application to facilitate the clinical interpretation of genetic and genomic variants from exome sequencing projects Raul Ossio MD1, Diego Said Anaya-Mancilla BSc1, O. Isaac Garcia-Salinas BSc1, Jair S. Garcia-Sotelo MSc1, Luis A. Aguilar MSc1, David J. Adams PhD2 and Carla Daniela Robles-Espinoza PhD1,2* 1Laboratorio Internacional de Investigación sobre el Genoma Humano, Universidad Nacional Autónoma de México, Querétaro, Qro., 76230, Mexico 2Experimental Cancer Genetics, Wellcome Sanger Institute, Hinxton Cambridge, CB10 1SA, UK * To whom correspondence should be addressed. Carla Daniela Robles-Espinoza Tel: +52 55 56 23 43 31 ext. 259 Email: [email protected] 1 bioRxiv preprint doi: https://doi.org/10.1101/466490; this version posted November 14, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. CONFLICT OF INTEREST No conflicts of interest. 2 bioRxiv preprint doi: https://doi.org/10.1101/466490; this version posted November 14, 2018. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. ABSTRACT Purpose To create a user-friendly web application that allows researchers, medical professionals and patients to easily and securely view, filter and interact with human exome sequencing data in the Variant Call Format (VCF). -
Infravec2 Open Research Data Management Plan
INFRAVEC2 OPEN RESEARCH DATA MANAGEMENT PLAN Authors: Andrea Crisanti, Gareth Maslen, Andy Yates, Paul Kersey, Alain Kohl, Clelia Supparo, Ken Vernick Date: 10th July 2020 Version: 3.0 Overview Infravec2 will align to Open Research Data, as follows: Data Types and Standards Infravec2 will generate a variety of data types, including molecular data types: genome sequence and assembly, structural annotation (gene models, repeats, other functional regions) and functional annotation (protein function assignment), variation data, and transcriptome data; arbovirus and malaria experimental infection data, linked to archived samples; and microbiome data (Operational Taxonomic Units), including natural virome composition. All data will be released according to the appropriate standards and formats for each data type. For example, DNA sequence will be released in FASTA format; variant calls in Variant Call Format; sequence alignments in BAM (Binary Alignment Map) and CRAM (Compressed Read Alignment Map) formats, etc. We will strongly encourage the organisation of linked data sets as Track Hubs, a mechanism for publishing a set of linked genomic data that aids data discovery, sharing, and selection for subsequent analysis. We will develop internal standards within the consortium to define minimal metadata that will accompany all data sets, following the template of the Minimal Information Standards for Biological and Biomedical Investigations (http://www.dcc.ac.uk/resources/metadata-standards/mibbi-minimum-information-biological- and-biomedical-investigations). Data Exploitation, Accessibility, Curation and Preservation All molecular data for which existing public data repositories exist will be submitted to such repositories on or before the publication of written manuscripts, with early release of data (i.e. -
Alternate-Locus Aware Variant Calling in Whole Genome Sequencing Marten Jäger1,2, Max Schubach1, Tomasz Zemojtel1,Knutreinert3, Deanna M
Jäger et al. Genome Medicine (2016) 8:130 DOI 10.1186/s13073-016-0383-z RESEARCH Open Access Alternate-locus aware variant calling in whole genome sequencing Marten Jäger1,2, Max Schubach1, Tomasz Zemojtel1,KnutReinert3, Deanna M. Church4 and Peter N. Robinson1,2,3,5,6* Abstract Background: The last two human genome assemblies have extended the previous linear golden-path paradigm of the human genome to a graph-like model to better represent regions with a high degree of structural variability. The new model offers opportunities to improve the technical validity of variant calling in whole-genome sequencing (WGS). Methods: We developed an algorithm that analyzes the patterns of variant calls in the 178 structurally variable regions of the GRCh38 genome assembly, and infers whether a given sample is most likely to contain sequences from the primary assembly, an alternate locus, or their heterozygous combination at each of these 178 regions. We investigate 121 in-house WGS datasets that have been aligned to the GRCh37 and GRCh38 assemblies. Results: We show that stretches of sequences that are largely but not entirely identical between the primary assembly and an alternate locus can result in multiple variant calls against regions of the primary assembly. In WGS analysis, this results in characteristic and recognizable patterns of variant calls at positions that we term alignable scaffold-discrepant positions (ASDPs). In 121 in-house genomes, on average 51.8 ± 3.8 of the 178 regions were found to correspond best to an alternate locus rather than the primary assembly sequence, and filtering these genomes with our algorithm led to the identification of 7863 variant calls per genome that colocalized with ASDPs. -
Galaxy Platform for NGS Data Analyses
Galaxy Platform For NGS Data Analyses Weihong Yan [email protected] Collaboratory Web Site http://qcb.ucla.edu/collaboratory Collaboratory Workshops Workshop Outline ü Day 1 § UCLA galaxy and user account § Galaxy web interface and management § Tools for NGS analyses and their application § Data formats § Build/share workflow and history § Q and A ü Day 2 § Galaxy Tools for RNA-seq analysis § Galaxy Tools for ChIP-seq analysis § Galaxy Tools for annotation. § Q and A *** Published datasets/results will be used in the tutorial UCLA Galaxy http://galaxy.hoffman2.idre.ucla.edu ü Hardware – Headnode (1) 96Gb memory, 12 core – Computing nodes (8) 48Gb memory, 12 core – Storage 100 Tb disk space ü Galaxy Resource Management - Hoffman2 grid engine Default: 1 core/job bowtie, bwa, tophat, cuffdiff, cufflinks, gatk programs: 4 core/job UCLA Galaxy http://galaxy.hoffman2.idre.ucla.edu ü galaxy login account: login: your email associated with ucla ü Disk quota: 1 Tb/user Galaxy Account Management Installed tools Launch analysis and view result History of execu7on and results Raw Reads *_qseq.txt, *.fastq Upload to Galaxy File transfer protocol (ftp) deMultiplex Barcode splitter, deMultiplex workflow fastqc, compute quality statistics, Quality Assessment draw quality score boxplot, draw nuclotides distribution Process Reads Trim sequences, sickle, scythe Alignment to bwa, bowtie, bowtie2, tophat Reference Format Conversion Text manipulation toolkit, BEDTools, SAM Results (sam/bam) Tools, java genomics toolkit, picard toolkit Downstream Analyses BS-Seeker2, cufflinks, cuffdiff, macs, macs2, GATK, CEAS Visualization Genome browser, IGV Repositories of Galaxy Tools https://toolshed.g2.bx.psu.edu ü History panel contains all datasets that are uploaded and results derived from certain analyses ü A history can be organized, annotated, and managed as a project ü History is sharable. -
Semantics of Data for Life Sciences and Reproducible Research[Version 1
F1000Research 2020, 9:136 Last updated: 16 AUG 2021 OPINION ARTICLE BioHackathon 2015: Semantics of data for life sciences and reproducible research [version 1; peer review: 2 approved] Rutger A. Vos 1,2, Toshiaki Katayama 3, Hiroyuki Mishima4, Shin Kawano 3, Shuichi Kawashima3, Jin-Dong Kim3, Yuki Moriya3, Toshiaki Tokimatsu5, Atsuko Yamaguchi 3, Yasunori Yamamoto3, Hongyan Wu6, Peter Amstutz7, Erick Antezana 8, Nobuyuki P. Aoki9, Kazuharu Arakawa10, Jerven T. Bolleman 11, Evan Bolton12, Raoul J. P. Bonnal13, Hidemasa Bono 3, Kees Burger14, Hirokazu Chiba15, Kevin B. Cohen16,17, Eric W. Deutsch18, Jesualdo T. Fernández-Breis19, Gang Fu12, Takatomo Fujisawa20, Atsushi Fukushima 21, Alexander García22, Naohisa Goto23, Tudor Groza24,25, Colin Hercus26, Robert Hoehndorf27, Kotone Itaya10, Nick Juty28, Takeshi Kawashima20, Jee-Hyub Kim28, Akira R. Kinjo29, Masaaki Kotera30, Kouji Kozaki 31, Sadahiro Kumagai32, Tatsuya Kushida 33, Thomas Lütteke 34,35, Masaaki Matsubara 36, Joe Miyamoto37, Attayeb Mohsen 38, Hiroshi Mori39, Yuki Naito3, Takeru Nakazato3, Jeremy Nguyen-Xuan40, Kozo Nishida41, Naoki Nishida42, Hiroyo Nishide15, Soichi Ogishima43, Tazro Ohta3, Shujiro Okuda44, Benedict Paten45, Jean-Luc Perret46, Philip Prathipati38, Pjotr Prins47,48, Núria Queralt-Rosinach 49, Daisuke Shinmachi9, Shinya Suzuki 30, Tsuyosi Tabata50, Terue Takatsuki51, Kieron Taylor 28, Mark Thompson52, Ikuo Uchiyama 15, Bruno Vieira53, Chih-Hsuan Wei12, Mark Wilkinson 54, Issaku Yamada 36, Ryota Yamanaka55, Kazutoshi Yoshitake56, Akiyasu C. Yoshizawa50, Michel -
Practical Guideline for Whole Genome Sequencing Disclosure
Practical Guideline for Whole Genome Sequencing Disclosure Kwangsik Nho Assistant Professor Center for Neuroimaging Department of Radiology and Imaging Sciences Center for Computational Biology and Bioinformatics Indiana University School of Medicine • Kwangsik Nho discloses that he has no relationships with commercial interests. What You Will Learn Today • Basic File Formats in WGS • Practical WGS Analysis Pipeline • WGS Association Analysis Methods Whole Genome Sequencing File Formats WGS Sequencer Base calling FASTQ: raw NGS reads Aligning SAM: aligned NGS reads BAM Variant Calling VCF: Genomic Variation How have BIG data problems been solved in next generation sequencing? gkno.me Whole Genome Sequencing File Formats • FASTQ: text-based format for storing both a DNA sequence and its corresponding quality scores (File sizes are huge (raw text) ~300GB per sample) @HS2000-306_201:6:1204:19922:79127/1 ACGTCTGGCCTAAAGCACTTTTTCTGAATTCCACCCCAGTCTGCCCTTCCTGAGTGCCTGGGCAGGGCCCTTGGGGAGCTGCTGGTGGGGCTCTGAATGT + BC@DFDFFHHHHHJJJIJJJJJJJJJJJJJJJJJJJJJHGGIGHJJJJJIJEGJHGIIJJIGGIIJHEHFBECEC@@D@BDDDDD<<?DB@BDDCDCDDC @HS2000-306_201:6:1204:19922:79127/2 GGGTAAAAGGTGTCCTCAGCTAATTCTCATTTCCTGGCTCTTGGCTAATTCTCATTCCCTGGGGGCTGGCAGAAGCCCCTCAAGGAAGATGGCTGGGGTC + BCCDFDFFHGHGHIJJJJJJJJJJJGJJJIIJCHIIJJIJIJJJJIJCHEHHIJJJJJJIHGBGIIHHHFFFDEEDED?B>BCCDDCBBDACDD8<?B09 @HS2000-306_201:4:1106:6297:92330/1 CACCATCCCGCGGAGCTGCCAGATTCTCGAGGTCACGGCTTACACTGCGGAGGGCCGGCAACCCCGCCTTTAATCTGCCTACCCAGGGAAGGAAAGCCTC + CCCFFFFFHGHHHJIJJJJJJJJIJJIIIGIJHIHHIJIGHHHHGEFFFDDDDDDDDDDD@ADBDDDDDDDDDDDDEDDDDDABCDDB?BDDBDCDDDDD -
Assembly, Annotation, and Polymorphic Characterization of the Erysiphe Necator Transcriptome
Rochester Institute of Technology RIT Scholar Works Theses 8-10-2012 Assembly, annotation, and polymorphic characterization of the Erysiphe necator transcriptome Jason Myers Follow this and additional works at: https://scholarworks.rit.edu/theses Recommended Citation Myers, Jason, "Assembly, annotation, and polymorphic characterization of the Erysiphe necator transcriptome" (2012). Thesis. Rochester Institute of Technology. Accessed from This Thesis is brought to you for free and open access by RIT Scholar Works. It has been accepted for inclusion in Theses by an authorized administrator of RIT Scholar Works. For more information, please contact [email protected]. Assembly, Annotation, and Polymorphic Characterization of the Erysiphe necator Transcriptome by Jason Myers Submitted in partial fulfillment of the requirements for the Master of Science degree in Bioinformatics at Rochester Institute of Technology. Department of Biological Sciences School of Life Sciences Rochester Institute of Technology Rochester, NY August 10, 2012 Committee: Dr. Gary Skuse Associate Head of the School Life Sciences/Professor/Committee Member Dr. Michael Osier Bioinformatics Program Head/Program Advisor/Associate Professor/Committee Member Dr. Dina Newman Thesis Advisor/Associate Professor Dr. Lance Cadle-Davidson Associate Thesis Advisor/Plant Pathologist USDA-ARS Dr. Angela Baldo Thesis Project Advisor/Computational Biologist USDA-ARS II Abstract: The objectives of this study were to develop a transcriptomic reference resource and to characterize polymorphism between isolates of Erysiphe necator (syn. Uncinula necator), grape powdery mildew. The wine and fresh fruit markets are economically vital to many countries worldwide, and E. necator infection can cause severe crop damage and subsequent financial loss. Most of the publicly available sequence data for Erysiphales are from research done on Blumeria graminis f. -
Biomolecule and Bioentity Interaction Databases in Systems Biology: a Comprehensive Review
biomolecules Review Biomolecule and Bioentity Interaction Databases in Systems Biology: A Comprehensive Review Fotis A. Baltoumas 1,* , Sofia Zafeiropoulou 1, Evangelos Karatzas 1 , Mikaela Koutrouli 1,2, Foteini Thanati 1, Kleanthi Voutsadaki 1 , Maria Gkonta 1, Joana Hotova 1, Ioannis Kasionis 1, Pantelis Hatzis 1,3 and Georgios A. Pavlopoulos 1,3,* 1 Institute for Fundamental Biomedical Research, Biomedical Sciences Research Center “Alexander Fleming”, 16672 Vari, Greece; zafeiropoulou@fleming.gr (S.Z.); karatzas@fleming.gr (E.K.); [email protected] (M.K.); [email protected] (F.T.); voutsadaki@fleming.gr (K.V.); [email protected] (M.G.); hotova@fleming.gr (J.H.); [email protected] (I.K.); hatzis@fleming.gr (P.H.) 2 Novo Nordisk Foundation Center for Protein Research, University of Copenhagen, 2200 Copenhagen, Denmark 3 Center for New Biotechnologies and Precision Medicine, School of Medicine, National and Kapodistrian University of Athens, 11527 Athens, Greece * Correspondence: baltoumas@fleming.gr (F.A.B.); pavlopoulos@fleming.gr (G.A.P.); Tel.: +30-210-965-6310 (G.A.P.) Abstract: Technological advances in high-throughput techniques have resulted in tremendous growth Citation: Baltoumas, F.A.; of complex biological datasets providing evidence regarding various biomolecular interactions. Zafeiropoulou, S.; Karatzas, E.; To cope with this data flood, computational approaches, web services, and databases have been Koutrouli, M.; Thanati, F.; Voutsadaki, implemented to deal with issues such as data integration, visualization, exploration, organization, K.; Gkonta, M.; Hotova, J.; Kasionis, scalability, and complexity. Nevertheless, as the number of such sets increases, it is becoming more I.; Hatzis, P.; et al. Biomolecule and and more difficult for an end user to know what the scope and focus of each repository is and how Bioentity Interaction Databases in redundant the information between them is.