Introduction to Next Generation Sequencing
|
|
- Lucas Campbell
- 5 years ago
- Views:
Transcription
1 The Sequencing Revolution Introduction to Next Generation Sequencing Dena Leshkowitz,WIS 1 st BIOmics Workshop High throughput Short Read Sequencing Technologies Highly parallel reactions (millions to billions possible) Performed on cloned DNA populations Companies 454/Roche - Launched in 2005 Pyrosequencing by synthesis Solexa/Illumina - Launched in late 2006 Reversible terminator sequencing by synthesis (dye labeled nucleotides) Agencourt/ABI/Invitrogen Launched mid-2007 Sequencing by ligation (dye labeled dinucleotides) DNA Sequencing Throughput History Cost for Sequencing the Human Genome Objectives Illumina Genome Analyzer pipeline overview Pipeline Components Overview Master Script (GOAT) Interpreting pipeline s output Tools and approaches to further analyze the sequences Image Analysis Base Calling Alignment GERALD
2 Technology Overview Image Analysis & Base Calling flow cell A flow cell contains eight lanes Lane 1... Lane 8 Each cluster at each cycle, generates 4 fluorescence intensities Each lane contains two columns, each column contains up to 50 tiles Column 1 Column 2 DNA clusters are located and quantified, across all images Each tile is imaged four times per cycle one image per base Naively: highest of the 4 values determines the base Pipeline Components Overview Master Script (GOAT) Base Calling: Intensity Correction Cross talk correction: emission spectra of the four dyes overlaps Normalization: scaling factor to make intensities equivalent Image Analysis Base Calling Alignment Emission spectra of dye X Y GERALD X Y Base Calling Phasing/Prephasing Correction Phasing Prephasing Base Calling G C C C C C A Corrected Intensity C quality score Requires a sample with a random, balanced base composition and therefore is usually done on Phix our control A C G T
3 Quality Score Pipeline Components Overview Each base has a quality score Solexa's base scoring is similar to Phred scoresa way of expressing estimates of sequencing error probabilities. Q phred = -10 log10( Pe ) Pe = error probability of a particular base call Q20 = 1 error in 100 bases Q30 = 1 error in 1000 bases Master Script (GOAT) Image Analysis Base Calling Alignment GERALD The quality score is in ASCII format ASCII character code= quality value + 64 Quality Filtering GERALD Chastity threshold: The ratio of the brightest intensity over the sum of the brightest and second brightest intensities I A C = >0.6 I + I A B Filter (pure-bases): I A sequence which has a B chastity less than 0.6 on two or more bases among the first 25 bases will be filtered I A ELAND Very fast Alignment: Program ELAND Only 2 mismatches allowed in first 32 bases (N is not counted as a mismatch) Alignments are used to estimate error rates Alignment: Programs Gerald (Eland) Objectives Eland Types Application Description Illumina Genome Analyzer pipeline overview Eland_extended Eland_pair Single reads Paired reads Aligns single reads to a reference Aligns paired reads Interpreting pipeline s output Eland_tag DGE Aligns to a nonredundant reference set of sequence tags Tools and approaches to further analyze the sequences Eland_rna Single reads, whole transcriptome Aligns to a reference genome, splice junctions and contaminations
4 Sequence Output Formats FASTQ (s_1_sequence.txt) Sequence Output Line 1: Unique ID for a sequencing read Line 2: Sequences Line 3: Repeat of the ID (preceded with a + sign) Line 4: Base calling quality score (Analogous to Phred scores but in ASCII value) ATTCCCCTGTACTGAGACATAGAGAGTTTGCAAGACCA +30LH2AAXX:8:1:984:225 \\\\\\\\\\Z\\\ZZZ\\\\\\W\\\\\ZYYYVYVVV Eland Alignment Outputs ELAND Outputs s_n_export.txt Results of alignment of all reads in the lane. The fields are tab separated to facilitate export to databases. The last field on each line is a flag telling you whether or not the read passed the filter (Y or N). s_n_sorted.txt Contains only entries for reads which : pass pure bases filtering have a unique alignment in the reference. Alignments are sorted by order of their alignment position Example : 30LL2AAXX ACGTGCTTACCCTACCACTCTATACCACCATCACTACC UUUUUUUUUUUUUUUUU UUUUUULUULUUUQQOQQIOO NC_ fna 354 F 19T10C3ATG1 0 30LL2AAXX ACGTGCTTACCCTACCACTTTATACCACCACCACATGC UUUUUUUUUUUUUUUUU UUUUUUUUUUUUUQQQQQOMO NC_ fna 354 F LL2AAXX TACCCTACCACTTTATACCACCACCACATGCCATACTC UUUUUUUUUUUUUUUUU Alignment File Format Tab Delimited Run Folder name Lane Tile X Coordinate of cluster Y Coordinate of cluster Index string (Blank for a non-indexed run) Read number (1 or 2 for paired-read analysis) Read Quality string In symbolic ASCII format Match chromosome Name of chromosome match OR code indicating why no match resulted Match Contig Gives the contig name Match Position Always with respect to forward strand Match Strand F for forward, R for reverse Match Descriptor Concise description of alignment Single-Read Alignment Score Paired-Read Alignment Score Partner Chromosome -paired read Partner Contig- paired read Partner Offset Partner Strand Filtering Did the read pass quality filtering? Y for yes, N for no 30LH2AAXX CAAATATGTTCAACAAAATTATAGTAGAAA GCTTTCCA ]]]]]]]]]]]]]]]]\]]]]]]\\]Z]]]YYYYYVVV NC_ fasta F 30A7 11 Y Run Statistics Quality Control
5 Summary.htm Report Folder Run Statistics Summary.htm (Report folder) The number of detected clusters The number of cluster that Passed Filtering The average intensity of all color channels in all tiles for the first cycle. Should be above 100 Percent intensity after 20 cycles should be 50% or more %PF should be above 50% (possible problems: too many clusters, faint clusters ) %Aligned filtered reads uniquely aligned %Error rate Should be 1.5 and below The percentage of each base called as a function of the cycle. Each channel (ATGC) is plotted separately IVC.htm Intensity Versus Cycle The red bar shows the % of bases at each cycle that are wrong, based on the eland alignment The error rate raises with the cycles Remark: the sequences were selected upon there ability to align to the first 32 bases Error.htm Pipeline Outputs You will find the following folders within the folder run: Folder Data type Folder structure: Storage Space GERALD_ FINAL_ Report Original folder from pipeline Original folder from pipeline Original Gerald folder (can contain CASAVA) from pipeline Final text outputs: Sequences Alignments Summarized as web page Summary.htm Optional data Optional data Optional data (also found in GERALD) (also found in GERALD) FC1012X Gerald Images 750Gb 250Gb Transferred to storage server (dapsas) <100Gb
6 Statistics of Runs Performed Objectives Illumina Genome Analyzer pipeline overview Interpreting pipeline s output Tools and approaches to further analyze the sequences The Jigsaw Puzzle One Run with 4Gb made of 100 million pieces each of length of 40 bases and some do not fit correctly. Mapping: Aligning to a reference sequence 1. Resequencing 2. Transcriptome analysis (RNA-seq) 3. Cistrome analysis (Chip-seq) First Step in Analysis Sequence data (bases & quality) De novo Assembly: Assembling individual sequences to a larger sequences De Novo Sequencing Example: Pseudomonas syringae Butler et al. FEMS Microbiol Lett 291 (2009) million genome X42 coverage ~3.5 million paired end reads of 36 bases De novo assembly using VELVET and EDENA, at least 3% of the reference genome was absent from the assembly (842 unassembled regions). Unassembled regions are noncoding RNA 90% of the protein-coding genes being assembled with 100% accuracy over their full length Differences Among the Mapping Applications Speed (Bowtie -string matching using Burrows Wheeler Transform) Use of quality data (MAQ, consed) Ability to perform multiple mapping (Nexalign) Amount of mismatches and indels supported (Soap) Length of seed alignment supported (Eland -32bases)
7 Resequencing Example SNP Detection & Reporting using CONSED Consed Can Detect Inserted Base CASAVA Consensus Assessment of Sequence And Variation (Illumina) RNA-Seq Post sequencing analysis: uses the export.txt files from the Eland alignment as input For resequencing projects: produces a set of allele calls of SNPs For RNA-seq (whole transcriptome sequencing): provides counts for exons, genes and splice junctions RNA-seq The expression value is calculated by counting the number of reads per gene, exon or splice junction Normalization of the expression value is done by: Dividing the number of reads by the virtual length of the gene or exon Scaling the number of reads between the samples RNA Seq An example of alternative splicing Chromosome Start End GeneSymbol Count_Normalized Lane2 Count_Lane2 c PRB c STATH c PIP c PRR c PRB c SLPI by Cold Spring Harbor Laboratory Press Marioni J C et al. Genome Res. 2008;18:
8 Basic output files: BED An example of Bed format file for reads that mapped to a genome: Visualize the Sequence Data Importing to Genome Viewers CHR: START: STOP: NAME: COUNT: STRAND: chr seqname 2 + chr seqname 3 + chr seqname 4 + chr seqname 26 + chr seqname 2 + chr seqname 3 + chr seqname 1 + Basic output files: WIG Sequencing "signal" - wiggle track: Imported Bed & Wiggle files to IGB genome browser Locus Signal variablestep chrom=chr Defining DNA protein interactions Chip-Seq MACS: Model-based Analysis for ChIP-Seq Binding Use confident peaks to model shift size Sultan et al. Science Aug 15;321(5891): CSHL Shirley Liu
9 Example of a Peak (MACS) Objectives Illumina Genome Analyzer pipeline overview Interpreting pipeline s output Tools and approaches to further analyze the sequences chr chr1 start end length 684 summit 278 tags 68-10LOG10 *(pvalue) Fold enrich ment FDR (%) 0.84 Bioinformatics wiki THANKS See you at the workshop this afternoon Everybody is invited to read and add to this wiki!
NEXT GENERATION SEQUENCING. Farhat Habib
NEXT GENERATION SEQUENCING HISTORY HISTORY Sanger Dominant for last ~30 years 1000bp longest read Based on primers so not good for repetitive or SNPs sites HISTORY Sanger Dominant for last ~30 years 1000bp
More informationLecture 7. Next-generation sequencing technologies
Lecture 7 Next-generation sequencing technologies Next-generation sequencing technologies General principles of short-read NGS Construct a library of fragments Generate clonal template populations Massively
More informationSequence Assembly and Alignment. Jim Noonan Department of Genetics
Sequence Assembly and Alignment Jim Noonan Department of Genetics james.noonan@yale.edu www.yale.edu/noonanlab The assembly problem >>10 9 sequencing reads 36 bp - 1 kb 3 Gb Outline Basic concepts in genome
More informationBioinformatics in next generation sequencing projects
Bioinformatics in next generation sequencing projects Rickard Sandberg Assistant Professor Department of Cell and Molecular Biology Karolinska Institutet May 2013 Standard sequence library generation Illumina
More informationBST 226 Statistical Methods for Bioinformatics David M. Rocke. March 10, 2014 BST 226 Statistical Methods for Bioinformatics 1
BST 226 Statistical Methods for Bioinformatics David M. Rocke March 10, 2014 BST 226 Statistical Methods for Bioinformatics 1 NGS Technologies Illumina Sequencing HiSeq 2500 & MiSeq PacBio Sequencing PacBio
More informationDeep Sequencing technologies
Deep Sequencing technologies Gabriela Salinas 30 October 2017 Transcriptome and Genome Analysis Laboratory http://www.uni-bc.gwdg.de/index.php?id=709 Microarray and Deep-Sequencing Core Facility University
More informationNext Generation Sequencing. Tobias Österlund
Next Generation Sequencing Tobias Österlund tobiaso@chalmers.se NGS part of the course Week 4 Friday 13/2 15.15-17.00 NGS lecture 1: Introduction to NGS, alignment, assembly Week 6 Thursday 26/2 08.00-09.45
More informationData Analysis with CASAVA v1.8 and the MiSeq Reporter
Data Analysis with CASAVA v1.8 and the MiSeq Reporter Eric Smith, PhD Bioinformatics Scientist September 15 th, 2011 2010 Illumina, Inc. All rights reserved. Illumina, illuminadx, Solexa, Making Sense
More informationMapping Next Generation Sequence Reads. Bingbing Yuan Dec. 2, 2010
Mapping Next Generation Sequence Reads Bingbing Yuan Dec. 2, 2010 1 What happen if reads are not mapped properly? Some data won t be used, thus fewer reads would be aligned. Reads are mapped to the wrong
More informationNext-generation sequencing and quality control: An introduction 2016
Next-generation sequencing and quality control: An introduction 2016 s.schmeier@massey.ac.nz http://sschmeier.com/bioinf-workshop/ Overview Typical workflow of a genomics experiment Genome versus transcriptome
More informationAbout Strand NGS. Strand Genomics, Inc All rights reserved.
About Strand NGS Strand NGS-formerly known as Avadis NGS, is an integrated platform that provides analysis, management and visualization tools for next-generation sequencing data. It supports extensive
More informationDe Novo Assembly of High-throughput Short Read Sequences
De Novo Assembly of High-throughput Short Read Sequences Chuming Chen Center for Bioinformatics and Computational Biology (CBCB) University of Delaware NECC Third Skate Genome Annotation Workshop May 23,
More informationAnalysing genomes and transcriptomes using Illumina sequencing
Analysing genomes and transcriptomes using Illumina uencing Dr. Heinz Himmelbauer Centre for Genomic Regulation (CRG) Ultrauencing Unit Barcelona The Sequencing Revolution High-Throughput Sequencing 2000
More informationNext Generation Sequencing: An Overview
Next Generation Sequencing: An Overview Cavan Reilly November 13, 2017 Table of contents Next generation sequencing NGS and microarrays Study design Quality assessment Burrows Wheeler transform Next generation
More informationServices Presentation Genomics Experts
Services Presentation Genomics Experts Illumina Seminar Marriott May 11th IntegraGen at a glance Autism Oncology Genomics Services Serves the researcher s most complex needs in genomics The n 1 privately-owned
More informationTranscriptomics analysis with RNA seq: an overview Frederik Coppens
Transcriptomics analysis with RNA seq: an overview Frederik Coppens Platforms Applications Analysis Quantification RNA content Platforms Platforms Short (few hundred bases) Long reads (multiple kilobases)
More informationIntroduction to RNA-Seq in GeneSpring NGS Software
Introduction to RNA-Seq in GeneSpring NGS Software Dipa Roy Choudhury, Ph.D. Strand Scientific Intelligence and Agilent Technologies Learn more at www.genespring.com Introduction to RNA-Seq In a few years,
More informationRead Quality Assessment & Improvement. UCD Genome Center Bioinformatics Core Tuesday 14 June 2016
Read Quality Assessment & Improvement UCD Genome Center Bioinformatics Core Tuesday 14 June 2016 QA&I should be interactive Error modes Each technology has unique error modes, depending on the physico-chemical
More informationData Basics. Josef K Vogt Slides by: Simon Rasmussen Next Generation Sequencing Analysis
Data Basics Josef K Vogt Slides by: Simon Rasmussen 2017 Generalized NGS analysis Sample prep & Sequencing Data size Main data reductive steps SNPs, genes, regions Application Assembly: Compare Raw Pre-
More informationNext Gen Sequencing. Expansion of sequencing technology. Contents
Next Gen Sequencing Contents 1 Expansion of sequencing technology 2 The Next Generation of Sequencing: High-Throughput Technologies 3 High Throughput Sequencing Applied to Genome Sequencing (TEDed CC BY-NC-ND
More informationBasics of RNA-Seq. (With a Focus on Application to Single Cell RNA-Seq) Michael Kelly, PhD Team Lead, NCI Single Cell Analysis Facility
2018 ABRF Meeting Satellite Workshop 4 Bridging the Gap: Isolation to Translation (Single Cell RNA-Seq) Sunday, April 22 Basics of RNA-Seq (With a Focus on Application to Single Cell RNA-Seq) Michael Kelly,
More informationThe New Genome Analyzer IIx Delivering more data, faster, and easier than ever before. Jeremy Preston, PhD Marketing Manager, Sequencing
The New Genome Analyzer IIx Delivering more data, faster, and easier than ever before Jeremy Preston, PhD Marketing Manager, Sequencing Illumina Genome Analyzer: a Paradigm Shift 2000x gain in efficiency
More informationMatthew Tinning Australian Genome Research Facility. July 2012
Next-Generation Sequencing: an overview of technologies and applications Matthew Tinning Australian Genome Research Facility July 2012 History of Sequencing Where have we been? 1869 Discovery of DNA 1909
More informationGenomic resources. for non-model systems
Genomic resources for non-model systems 1 Genomic resources Whole genome sequencing reference genome sequence comparisons across species identify signatures of natural selection population-level resequencing
More informationRADseq Data Analysis Workshop 3 February 2017
RADseq Data Analysis Workshop 3 February 2017 Introduction to Galaxy (thanks to Simon Gladman for slides) What is Galaxy? A web-based scalable workflow platform for genomic analysis Designed for biologists
More informationRNAseq Applications in Genome Studies. Alexander Kanapin, PhD Wellcome Trust Centre for Human Genetics, University of Oxford
RNAseq Applications in Genome Studies Alexander Kanapin, PhD Wellcome Trust Centre for Human Genetics, University of Oxford RNAseq Protocols Next generation sequencing protocol cdna, not RNA sequencing
More informationGenome 373: Mapping Short Sequence Reads II. Doug Fowler
Genome 373: Mapping Short Sequence Reads II Doug Fowler The final Will be in this room on June 6 th at 8:30a Will be focused on the second half of the course, but will include material from the first half
More information02 Agenda Item 03 Agenda Item
01 Agenda Item 02 Agenda Item 03 Agenda Item SOLiD 3 System: Applications Overview April 12th, 2010 Jennifer Stover Field Application Specialist - SOLiD Applications Workflow for SOLiD Application Application
More informationThe Expanded Illumina Sequencing Portfolio New Sample Prep Solutions and Workflow
The Expanded Illumina Sequencing Portfolio New Sample Prep Solutions and Workflow Marcus Hausch, Ph.D. 2010 Illumina, Inc. All rights reserved. Illumina, illuminadx, Solexa, Making Sense Out of Life, Oligator,
More informationGalaxy for Next Generation Sequencing 初探次世代序列分析平台 蘇聖堯 2013/9/12
Galaxy for Next Generation Sequencing 初探次世代序列分析平台 蘇聖堯 2013/9/12 What s Galaxy? Bringing Developers And Biologists Together. Reproducible Science Is Our Goal An open, web-based platform for data intensive
More informationReference genomes and common file formats
Reference genomes and common file formats Overview Reference genomes and GRC Fasta and FastQ (unaligned sequences) SAM/BAM (aligned sequences) Summarized genomic features BED (genomic intervals) GFF/GTF
More informationChIP-seq data analysis with Chipster. Eija Korpelainen CSC IT Center for Science, Finland
ChIP-seq data analysis with Chipster Eija Korpelainen CSC IT Center for Science, Finland chipster@csc.fi What will I learn? Short introduction to ChIP-seq Analyzing ChIP-seq data Central concepts Analysis
More informationReference genomes and common file formats
Reference genomes and common file formats Dóra Bihary MRC Cancer Unit, University of Cambridge CRUK Functional Genomics Workshop September 2017 Overview Reference genomes and GRC Fasta and FastQ (unaligned
More informationRNA-Sequencing analysis
RNA-Sequencing analysis Markus Kreuz 25. 04. 2012 Institut für Medizinische Informatik, Statistik und Epidemiologie Content: Biological background Overview transcriptomics RNA-Seq RNA-Seq technology Challenges
More informationAn introduction to RNA-seq. Nicole Cloonan - 4 th July 2018 #UQWinterSchool #Bioinformatics #GroupTherapy
An introduction to RNA-seq Nicole Cloonan - 4 th July 2018 #UQWinterSchool #Bioinformatics #GroupTherapy The central dogma Genome = all DNA in an organism (genotype) Transcriptome = all RNA (molecular
More informationNext Generation Sequencing Technologies. Some slides are modified from Robi Mitra s lecture notes
Next Generation Sequencing Technologies Some slides are modified from Robi Mitra s lecture notes What will you do to understand a disease? What will you do to understand a disease? Genotype Phenotype Hypothesis
More informationNext-Generation Sequencing. Technologies
Next-Generation Next-Generation Sequencing Technologies Sequencing Technologies Nicholas E. Navin, Ph.D. MD Anderson Cancer Center Dept. Genetics Dept. Bioinformatics Introduction to Bioinformatics GS011062
More informationContact us for more information and a quotation
GenePool Information Sheet #1 Installed Sequencing Technologies in the GenePool The GenePool offers sequencing service on three platforms: Sanger (dideoxy) sequencing on ABI 3730 instruments Illumina SOLEXA
More informationNGS in Pathology Webinar
NGS in Pathology Webinar NGS Data Analysis March 10 2016 1 Topics for today s presentation 2 Introduction Next Generation Sequencing (NGS) is becoming a common and versatile tool for biological and medical
More informationQIAseq Targeted Panel Analysis Plugin USER MANUAL
QIAseq Targeted Panel Analysis Plugin USER MANUAL User manual for QIAseq Targeted Panel Analysis 1.1 Windows, macos and Linux June 18, 2018 This software is for research purposes only. QIAGEN Aarhus Silkeborgvej
More informationBioinformatics small variants Data Analysis. Guidelines. genomescan.nl
Next Generation Sequencing Bioinformatics small variants Data Analysis Guidelines genomescan.nl GenomeScan s Guidelines for Small Variant Analysis on NGS Data Using our own proprietary data analysis pipelines
More informationIntroduction to bioinformatics (NGS data analysis)
Introduction to bioinformatics (NGS data analysis) Alexander Jueterbock 2015-06-02 1 / 45 Got your sequencing data - now, what to do with it? File size: several Gb Number of lines: >1,000,000 @M02443:17:000000000-ABPBW:1:1101:12675:1533
More informationChapter 7. DNA Microarrays
Bioinformatics III Structural Bioinformatics and Genome Analysis Chapter 7. DNA Microarrays 7.9 Next Generation Sequencing 454 Sequencing Solexa Illumina Solid TM System Sequencing Process of determining
More informationBioinformatics for NGS projects. Guidelines. genomescan.nl
Next Generation Sequencing Bioinformatics for NGS projects Guidelines genomescan.nl GenomeScan s Guidelines for Bioinformatics Services on NGS Data Using our own proprietary data analysis pipelines Dear
More informationReads to Discovery. Visualize Annotate Discover. Small DNA-Seq ChIP-Seq Methyl-Seq. MeDIP-Seq. RNA-Seq. RNA-Seq.
Reads to Discovery RNA-Seq Small DNA-Seq ChIP-Seq Methyl-Seq RNA-Seq MeDIP-Seq www.strand-ngs.com Analyze Visualize Annotate Discover Data Import Alignment Vendor Platforms: Illumina Ion Torrent Roche
More informationExperimental Design. Sequencing. Data Quality Control. Read mapping. Differential Expression analysis
-Seq Analysis Quality Control checks Reproducibility Reliability -seq vs Microarray Higher sensitivity and dynamic range Lower technical variation Available for all species Novel transcript identification
More information1. Introduction Gene regulation Genomics and genome analyses
1. Introduction Gene regulation Genomics and genome analyses 2. Gene regulation tools and methods Regulatory sequences and motif discovery TF binding sites Databases 3. Technologies Microarrays Deep sequencing
More informationRNA-Seq. Joshua Ainsley, PhD Postdoctoral Researcher Lab of Leon Reijmers Neuroscience Department Tufts University
RNA-Seq Joshua Ainsley, PhD Postdoctoral Researcher Lab of Leon Reijmers Neuroscience Department Tufts University joshua.ainsley@tufts.edu Day five Alternative splicing Assembly RNA edits Alternative splicing
More informationIntroduction to transcriptome analysis using High Throughput Sequencing technologies. D. Puthier 2012
Introduction to transcriptome analysis using High Throughput Sequencing technologies D. Puthier 2012 A typical RNA-Seq experiment Library construction Protocol variations Fragmentation methods RNA: nebulization,
More informationresequencing storage SNP ncrna metagenomics private trio de novo exome ncrna RNA DNA bioinformatics RNA-seq comparative genomics
RNA Sequencing T TM variation genetics validation SNP ncrna metagenomics private trio de novo exome mendelian ChIP-seq RNA DNA bioinformatics custom target high-throughput resequencing storage ncrna comparative
More informationWhole Transcriptome Analysis of Illumina RNA- Seq Data. Ryan Peters Field Application Specialist
Whole Transcriptome Analysis of Illumina RNA- Seq Data Ryan Peters Field Application Specialist Partek GS in your NGS Pipeline Your Start-to-Finish Solution for Analysis of Next Generation Sequencing Data
More informationThe first thing you will see is the opening page. SeqMonk scans your copy and make sure everything is in order, indicated by the green check marks.
Open Seqmonk Launch SeqMonk The first thing you will see is the opening page. SeqMonk scans your copy and make sure everything is in order, indicated by the green check marks. SeqMonk Analysis Page 1 Create
More informationSequencing technologies. Jose Blanca COMAV institute bioinf.comav.upv.es
Sequencing technologies Jose Blanca COMAV institute bioinf.comav.upv.es Outline Sequencing technologies: Sanger 2nd generation sequencing: 3er generation sequencing: 454 Illumina SOLiD Ion Torrent PacBio
More informationGalaxy Platform For NGS Data Analyses
Galaxy Platform For NGS Data Analyses Weihong Yan wyan@chem.ucla.edu Collaboratory Web Site http://qcb.ucla.edu/collaboratory http://collaboratory.lifesci.ucla.edu Workshop Outline ü Day 1 UCLA galaxy
More informationHow much sequencing do I need? Emily Crisovan Genomics Core
How much sequencing do I need? Emily Crisovan Genomics Core How much sequencing? Three questions: 1. How much sequence is required for good experimental design? 2. What type of sequencing run is best?
More informationChIP-seq and RNA-seq. Farhat Habib
ChIP-seq and RNA-seq Farhat Habib fhabib@iiserpune.ac.in Biological Goals Learn how genomes encode the diverse patterns of gene expression that define each cell type and state. Protein-DNA interactions
More informationSequencing technologies. Jose Blanca COMAV institute bioinf.comav.upv.es
Sequencing technologies Jose Blanca COMAV institute bioinf.comav.upv.es Outline Sequencing technologies: Sanger 2nd generation sequencing: 3er generation sequencing: 454 Illumina SOLiD Ion Torrent PacBio
More informationGenomics AGRY Michael Gribskov Hock 331
Genomics AGRY 60000 Michael Gribskov gribskov@purdue.edu Hock 331 Computing Essentials Resources In this course we will assemble and annotate both genomic and transcriptomic sequence assemblies We will
More informationAlignment methods. Martijn Vermaat Department of Human Genetics Center for Human and Clinical Genetics
Alignment methods Martijn Vermaat Department of Human Genetics Center for Human and Clinical Genetics Alignment methods Sequence alignment Assembly vs alignment Alignment methods Common issues Platform
More informationHigh Throughput Sequencing the Multi-Tool of Life Sciences. Lutz Froenicke DNA Technologies and Expression Analysis Cores UCD Genome Center
High Throughput Sequencing the Multi-Tool of Life Sciences Lutz Froenicke DNA Technologies and Expression Analysis Cores UCD Genome Center Complementary Approaches Illumina Still-imaging of clusters (~1000
More informationNature Methods: doi: /nmeth Supplementary Figure 1. Construction of a sensitive TetR mediated auxotrophic off-switch.
Supplementary Figure 1 Construction of a sensitive TetR mediated auxotrophic off-switch. A Production of the Tet repressor in yeast when conjugated to either the LexA4 or LexA8 promoter DNA binding sequences.
More informationSystematic evaluation of spliced alignment programs for RNA- seq data
Systematic evaluation of spliced alignment programs for RNA- seq data Pär G. Engström, Tamara Steijger, Botond Sipos, Gregory R. Grant, André Kahles, RGASP Consortium, Gunnar Rätsch, Nick Goldman, Tim
More informationIntroduction to RNA-Seq. David Wood Winter School in Mathematics and Computational Biology July 1, 2013
Introduction to RNA-Seq David Wood Winter School in Mathematics and Computational Biology July 1, 2013 Abundance RNA is... Diverse Dynamic Central DNA rrna Epigenetics trna RNA mrna Time Protein Abundance
More informationFast, Accurate and Sensitive DNA Variant Detection from Sanger Sequencing:
Fast, Accurate and Sensitive DNA Variant Detection from Sanger Sequencing: Patented, Anti-Correlation Technology Provides 99.5% Accuracy & Sensitivity to 5% Variant Knowledge Base and External Annotation
More informationIntroduction to human genomics and genome informatics
Introduction to human genomics and genome informatics Session 1 Prince of Wales Clinical School Dr Jason Wong ARC Future Fellow Head, Bioinformatics & Integrative Genomics Adult Cancer Program, Lowy Cancer
More informationRead Mapping and Variant Calling. Johannes Starlinger
Read Mapping and Variant Calling Johannes Starlinger Application Scenario: Personalized Cancer Therapy Different mutations require different therapy Collins, Meredith A., and Marina Pasca di Magliano.
More informationGene Expression analysis with RNA-Seq data
Gene Expression analysis with RNA-Seq data C3BI Hands-on NGS course November 24th 2016 Frédéric Lemoine Plan 1. 2. Quality Control 3. Read Mapping 4. Gene Expression Analysis 5. Splicing/Transcript Analysis
More informationIntroduction to the MiSeq
Introduction to the MiSeq 2011 Illumina, Inc. All rights reserved. Illumina, illuminadx, BeadArray, BeadXpress, cbot, CSPro, DASL, Eco, Genetic Energy, GAIIx, Genome Analyzer, GenomeStudio, GoldenGate,
More informationSequencing technologies. Jose Blanca COMAV institute bioinf.comav.upv.es
Sequencing technologies Jose Blanca COMAV institute bioinf.comav.upv.es Outline Sequencing technologies: Sanger 2nd generation sequencing: 3er generation sequencing: 454 Illumina SOLiD Ion Torrent PacBio
More informationAnalysis of neo-antigens to identify T-cell neo-epitopes in human Head & Neck cancer. Project XX1001. Customer Detail
Analysis of neo-antigens to identify T-cell neo-epitopes in human Head & Neck cancer Project XX Customer Detail Table of Contents. Bioinformatics analysis pipeline...3.. Read quality check. 3.2. Read alignment...3.3.
More information10/06/2014. RNA-Seq analysis. With reference assembly. Cormier Alexandre, PhD student UMR8227, Algal Genetics Group
RNA-Seq analysis With reference assembly Cormier Alexandre, PhD student UMR8227, Algal Genetics Group Summary 2 Typical RNA-seq workflow Introduction Reference genome Reference transcriptome Reference
More informationIllumina (Solexa) Throughput: 4 Tbp in one run (5 days) Cheapest sequencing technology. Mismatch errors dominate. Cost: ~$1000 per human genme
Illumina (Solexa) Current market leader Based on sequencing by synthesis Current read length 100-150bp Paired-end easy, longer matepairs harder Error ~0.1% Mismatch errors dominate Throughput: 4 Tbp in
More informationApplications of short-read
Applications of short-read sequencing: RNA-Seq and ChIP-Seq BaRC Hot Topics March 2013 George Bell, Ph.D. http://jura.wi.mit.edu/bio/education/hot_topics/ Sequencing applications RNA-Seq includes experiments
More informationTranscriptome analysis
Statistical Bioinformatics: Transcriptome analysis Stefan Seemann seemann@rth.dk University of Copenhagen April 11th 2018 Outline: a) How to assess the quality of sequencing reads? b) How to normalize
More informationQuantifying gene expression
Quantifying gene expression Genome GTF (annotation)? Sequence reads FASTQ FASTQ (+reference transcriptome index) Quality control FASTQ Alignment to Genome: HISAT2, STAR (+reference genome index) (known
More informationSUPPLEMENTARY INFORMATION
doi:1.138/nature11233 Supplementary Figure S1 Sample Flowchart. The ENCODE transcriptome data are obtained from several cell lines which have been cultured in replicates. They were either left intact (whole
More informationGene Expression Technology
Gene Expression Technology Bing Zhang Department of Biomedical Informatics Vanderbilt University bing.zhang@vanderbilt.edu Gene expression Gene expression is the process by which information from a gene
More informationAssembling a Cassava Transcriptome using Galaxy on a High Performance Computing Cluster
Assembling a Cassava Transcriptome using Galaxy on a High Performance Computing Cluster Aobakwe Matshidiso Supervisor: Prof Chrissie Rey Co-Supervisor: Prof Scott Hazelhurst Next Generation Sequencing
More informationFrom Variants to Pathways: Agilent GeneSpring GX s Variant Analysis Workflow
From Variants to Pathways: Agilent GeneSpring GX s Variant Analysis Workflow Technical Overview Import VCF Introduction Next-generation sequencing (NGS) studies have created unanticipated challenges with
More informationDNA concentration and purity were initially measured by NanoDrop 2000 and verified on Qubit 2.0 Fluorometer.
DNA Preparation and QC Extraction DNA was extracted from whole blood or flash frozen post-mortem tissue using a DNA mini kit (QIAmp #51104 and QIAmp#51404, respectively) following the manufacturer s recommendations.
More informationHow much sequencing do I need? Emily Crisovan Genomics Core September 26, 2018
How much sequencing do I need? Emily Crisovan Genomics Core September 26, 2018 How much sequencing? Three questions: 1. How much sequence is required for good experimental design? 2. What type of sequencing
More informationIntroduction to Bioinformatics and Gene Expression Technologies
Introduction to Bioinformatics and Gene Expression Technologies Utah State University Fall 2017 Statistical Bioinformatics (Biomedical Big Data) Notes 1 1 Vocabulary Gene: hereditary DNA sequence at a
More informationIntroduction to Bioinformatics and Gene Expression Technologies
Vocabulary Introduction to Bioinformatics and Gene Expression Technologies Utah State University Fall 2017 Statistical Bioinformatics (Biomedical Big Data) Notes 1 Gene: Genetics: Genome: Genomics: hereditary
More informationNext Generation Sequencing Lecture Saarbrücken, 19. March Sequencing Platforms
Next Generation Sequencing Lecture Saarbrücken, 19. March 2012 Sequencing Platforms Contents Introduction Sequencing Workflow Platforms Roche 454 ABI SOLiD Illumina Genome Anlayzer / HiSeq Problems Quality
More informationAnalysis of ChIP-seq data with R / Bioconductor
Analysis of ChIP-seq data with R / Bioconductor Martin Morgan Bioconductor / Fred Hutchinson Cancer Research Center Seattle, WA, USA 8-10 June 2009 ChIP-seq Chromatin immunopreciptation to enrich sample
More informationMODULE 5: TRANSLATION
MODULE 5: TRANSLATION Lesson Plan: CARINA ENDRES HOWELL, LEOCADIA PALIULIS Title Translation Objectives Determine the codons for specific amino acids and identify reading frames by looking at the Base
More informationWheat CAP Gene Expression with RNA-Seq
Wheat CAP Gene Expression with RNA-Seq July 9 th -13 th, 2018 Overview of the workshop, Alina Akhunova http://www.ksre.k-state.edu/igenomics/workshops/ RNA-Seq Workshop Activities Lectures Laboratory Molecular
More informationISO/IEC JTC 1/SC 29/WG 11 N15527 Warsaw, CH June Introduction
INTERNATIONAL ORGANISATION FOR STANDARDISATION ORGANISATION INTERNATIONALE DE NORMALISATION ISO/IEC JTC 1/SC 29/WG 11 CODING OF MOVING PICTURES AND AUDIO ISO/IEC JTC 1/SC 29/WG 11 N15527 Warsaw, CH June
More informationIntroductory Next Gen Workshop
Introductory Next Gen Workshop http://www.illumina.ucr.edu/ http://www.genomics.ucr.edu/ Workshop Objectives Workshop aimed at those who are new to Illumina sequencing and will provide: - a basic overview
More informationSequencing applications. Today's outline. Hands-on exercises. Applications of short-read sequencing: RNA-Seq and ChIP-Seq
Sequencing applications Applications of short-read sequencing: RNA-Seq and ChIP-Seq BaRC Hot Topics March 2013 George Bell, Ph.D. http://jura.wi.mit.edu/bio/education/hot_topics/ RNA-Seq includes experiments
More informationGenomic DNA ASSEMBLY BY REMAPPING. Course overview
ASSEMBLY BY REMAPPING Laurent Falquet, The Bioinformatics Unravelling Group, UNIFR & SIB MA/MER @ UniFr Group Leader @ SIB Course overview Genomic DNA PacBio Illumina methylation de novo remapping Annotation
More informationMapping strategies for sequence reads
Mapping strategies for sequence reads Ernest Turro University of Cambridge 21 Oct 2013 Quantification A basic aim in genomics is working out the contents of a biological sample. 1. What distinct elements
More informationA shotgun introduction to sequence assembly (with Velvet) MCB Brem, Eisen and Pachter
A shotgun introduction to sequence assembly (with Velvet) MCB 247 - Brem, Eisen and Pachter Hot off the press January 27, 2009 06:00 AM Eastern Time llumina Launches Suite of Next-Generation Sequencing
More informationIntroduction to NGS analyses
Introduction to NGS analyses Giorgio L Papadopoulos Institute of Molecular Biology and Biotechnology Bioinformatics Support Group 04/12/2015 Papadopoulos GL (IMBB, FORTH) IMBB NGS Seminar 04/12/2015 1
More informationWelcome to the NGS webinar series
Welcome to the NGS webinar series Webinar 1 NGS: Introduction to technology, and applications NGS Technology Webinar 2 Targeted NGS for Cancer Research NGS in cancer Webinar 3 NGS: Data analysis for genetic
More informationSupplement to: The Genomic Sequence of the Chinese Hamster Ovary (CHO)-K1 cell line
Supplement to: The Genomic Sequence of the Chinese Hamster Ovary (CHO)-K1 cell line Table of Contents SUPPLEMENTARY TEXT:... 2 FILTERING OF RAW READS PRIOR TO ASSEMBLY:... 2 COMPARATIVE ANALYSIS... 2 IMMUNOGENIC
More informationSequence Annotation & Designing Gene-specific qpcr Primers (computational)
James Madison University From the SelectedWorks of Ray Enke Ph.D. Fall October 31, 2016 Sequence Annotation & Designing Gene-specific qpcr Primers (computational) Raymond A Enke This work is licensed under
More informationRNA-Seq data analysis course September 7-9, 2015
RNA-Seq data analysis course September 7-9, 2015 Peter-Bram t Hoen (LUMC) Jan Oosting (LUMC) Celia van Gelder, Jacintha Valk (BioSB) Anita Remmelzwaal (LUMC) Expression profiling DNA mrna protein Comprehensive
More informationHigh-Throughput Bioinformatics: Re-sequencing and de novo assembly. Elena Czeizler
High-Throughput Bioinformatics: Re-sequencing and de novo assembly Elena Czeizler 13.11.2015 Sequencing data Current sequencing technologies produce large amounts of data: short reads The outputted sequences
More information