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Page 1: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

RNA-Seq Analysis

Page 2: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

QualityControlchecks• Reproducibility• Reliability

Page 3: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 4: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

RNA-seq vsMicroarray

• Highersensitivityanddynamicrange• Lowertechnicalvariation• Availableforallspecies• Noveltranscriptidentification• Alternatesplicing• Allelespecificexpression• Fusiongenes• HigherInformaticsCost

Page 5: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Reproducibility,LinearityandSensitivity

<2%ofgenome

Mortazavi etal.,NatureMethods5,621- 628(2008)

Page 6: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

RNAisolation

TotalRNA

Poly(A)selection

rRNA depletion SizeselectionClinicalSamples(tissuebiopsies)BacterialRNAsamples

SmallncRNA

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 7: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 8: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

GeneA GeneB GeneC

Sample1 6 1 4

GeneA GeneB GeneC Total

Sample1 3 1 1 5

Sample2 6 3 6 15

#ofreads

GeneA GeneB GeneC

Sample1 3 1 1Readsperkbofexon

GeneA GeneB GeneC Total

Sample1 0.6 0.2 0.2 5

Sample2 0.4 0.2 0.4 15

Readsperkbofexon

Readsperkbofexonpermillionmappedreads- RPKM

Page 9: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 10: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ExperimentalDesign

Whataremygoals?• Transcriptassembly• DifferentialExpressionanalysis• Identifynew/raretranscripts

WhatareCharacteristicsofmysystem?• Largeandcomplexgenome• Intronsandhighdegreeofalternativesplicing• Noreferencegenomeortranscriptome.

Page 11: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ExperimentalDesign

• BiologicalComparison(s)• PairedEndvsSingleend• Readdepth• Readlength• Replicates

Page 12: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ExperimentalDesign

SimpleDesign- Pairwisecomparison

TwoGroups

ComplexdesignControl Experimental

treatment

CancerSubtypeA

- +drug

CancerSubtypeB

- +drug

Consultastatistician

Page 13: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ExperimentalDesign

ReaddepthandreadlengthSmallgenomewithnoalternatesplicing(yeast)Annotatedtranscriptome

10millionreadspersample,50bpsingle-endreads

Mammaliangenomes(Largetranscriptome,alternativesplicing,geneduplication)

30millionreadspersample,

Transcriptomeassembly(100Xcoverageoftranscriptome)50-200millionreadspersample,100bppairedendreads

Natureofsamples.• Whatistheexpectedpurityofyoursample?• Istherecontaminationorheterogeneityexpected?

Ifyes,thenHighcoveragetodetectvariantsatlowerfrequencyduetoimpurityorbecausetheycomefromminor(butpossiblystillinteresting)subpopulationsofyoursample.

Page 14: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ExperimentalDesign

Replicates:Factorsdeterminingnumberofreplicates:• Variabilityinmeasurements(TechnicalnoiseandBiologicalvariation)• Statisticalpoweranalysis

TechnicalReplicatesNotNeeded:HighreproducibilityatsequencingstepErrorpronestepsRNAfragmentation,cDNAsynthesis,adapterligation,PCRamplification,bar-coding,laneloadingSpikeIns:Qualitycontrolandlibrary-sizenormalisationMinimizebatcheffectsRandomizesamplesatlibrarypreparationandsequencingruns

BiologicalReplicatesNotrequiredfortranscriptionassemblyEssentialfordifferentialexpressionanalysisComplexdesigns:

• 3+forcelllines• 5+forinbredlines• 20+forhumansamples

Page 15: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ExperimentalDesign

Scotty:http://scotty.genetics.utah.edu/

Page 16: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 17: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Sequencing

Illuminasequencingbysynthesis

SOLID“Color-Space”readsLowerrorrate

454pyrosequencingLongerreads,lowthroughput

Pacific-Bioscience(pacBio)/OxfordNanoporeLongerread(Recoveryoffulllengthtranscripts)

Page 18: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

Transcriptomeassembly

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

Page 19: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

Transcriptomeassembly

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

SequenceDataFormat.FASTQ

QualityScorePhred +33

ReadIDSequence

MachineID

QCFilterflagY=badN=goodReadpair#1

Readpair#2

Sample1_R1.fastqSample1_R2.fastq

R1

R2

SampleID/Barcode

Page 20: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

Evaluaterawreadlibraryquality• Sequencequality• GCcontentforbiases• AdapterContamination• K-mer overrepresentation• Duplicatereads• PCRartifacts

Software/Tools• FASTQC(Commandline)

Illuminareadfiles• NGSQC

SupportreadsfromanyplatformSupportqualitybasedreadtrimmingandfiltering

• SAMSat (Commandline)AlsoworkwithBamalignmentfiles

Page 21: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

Page 22: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

Sequencequality:Qualityscoresoverbases

Good

BadTrimmingrequired

Software/ToolsFastX-ToolkitTrimmomaticSickleCutadapt

Phred 30=1error/1000basesPhred 20=1error/100bases

Page 23: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

GCDistribution:AcceptablelevelsdependonSourceofsample

Good

Bad

Page 24: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

Page 25: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

Page 26: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DataQualityControl

DataQualityAssessment

Recommendations:

• Generatequalityplotsforallreadlibraries• Trimand/orfilterdataifneeded

Alwaystrimandfilterfordenovotranscriptomeassembly

• Regeneratequalityplotsaftertrimmingandfilteringtodetermineeffectiveness

• AcceptableduplicationK-mer orGCcontentlevelsareexperimentandorganismspecificbutthevaluesshouldbehomogeneousforsamplesinthesameexperiment.

• Outlierswith>30%disagreementshouldbediscarded.

Page 27: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 28: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ReadMapping

(1) Withreferencegenome(with/withouttranscriptome).(2) Withreferencetranscriptome.(3) Referencefreeassembly.

Page 29: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

RNA-seq:AssemblyvsMapping

RNA-seqreads

ReferencebasedRNA-seq

Ref:GenomeorTranscriptome

DenovoRNA-seqcontig 1 contig 2

Page 30: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Mappingwithreferencegenome

Shortreadsfastq

Mappingtogenome

Gappedmapper TopHatSTAR,HISAT2

Cufflinks,htseq-count,subread

Transcriptidentification&counting

WithGFF WithoutGFF

Transcriptdiscovery&counting

Functionalannotation

Blast2GOHomologybased

Page 31: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

MappingwithreferenceTranscriptome

Shortreadsfastq

MappingtoTranscriptome

Ungapped mapper Bowtie2

RSEMKallistoexpress

Transcriptidentification&counting

Page 32: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Mappingwithoutreference

Shortreadsfastq

Assemblyintotranscripts

DeBrujin graphs TrinityOasesVelvet

Mapreadsback

Ungapped mapper Bowtie

Htseq-countRSEM,express

Functionalannotation

GTF-based

Counting

Blast2GOHomologybased

Page 33: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Alignmenttools

Page 34: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Alignmenttools

Conesaetal,GenomeBiology201617:13

Page 35: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

`

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

ReadMapping

Page 36: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 37: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

StatisticalpropertiesofedgeR (exact)asafunctionoflog2(FC)threshold,T,andthenumberofreplicates,n

Page 38: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

DifferentialexpressionCommontoolsfordifferentialexpressionanalysis

Page 39: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Decisiontreeforsoftwareselection

Introduction

ExperimentalDesign

Sequencing

DataQualityControl

Readmapping

ReferenceGenome

ReferenceTranscriptome

DifferentialExpressionanalysis

RNA

fastq

fastq

SAM/BAM

fasta

GFF/GTF

Page 40: RNA seq Data Therapy 12Jan2018 - University of Connecticut · analysis RNA fastq fastq SAM/BAM fasta GFF/GTF RNA-seqvs Microarray • Higher sensitivity and dynamic range • Lower

Sources:

Conesaetal,GenomeBiology201617:13

Schurch etal,https://arxiv.org/abs/1505.02017

ZengAndMortazavi NatImmunol 13(9),802-7

Mortazavi etalNatureMethods volume5, pages621–628 (2008)


Galaxy environment Dr. Orr Shomroni Analysis of RNA ... · Per-base quality information (Phred score 33) RNA-seq workflow II – FASTQ processing Steps towards identifying differential

TRANSCRIPTION RNA is transcribed from a DNA template. DNA RNA polymerase RNA transcript RNA PROCESSING In eukaryotes, the RNA transcript (pre- mRNA) is

RNA-Regulationsmechanismen: RNA Interferenz · RNA Interferenz (Fire&Mello) RNA Moleküle können interferieren, d.h. insbesondere kann doppelsträngige RNA den Abbau von RNA und

From FastQ Data to High-Confidence Variant Calls: The Genome Analysis Toolkit Best Practices Pipeline

BWAWholeGenomeSequencing v1.0 BaseSpaceAppGuide · 2018-09-11 · Introduction BWAWholeGenomeSequencingv1.0BaseSpaceAppGuide 3 Introduction AfterBaseSpace®generates the FASTQ files

Turbo-Decoding of RNA Secondary Structure€¦ · RNA Structure Analysis: Motivation and Background RNA, noncoding RNA, RNA structure and its significance RNA structure prediction

SK 6L80 12Jan2018

CASAVA でつくる FASTQ - イルミナ株式会社 | 遺 … HiSeq/MiSeq データをFASTQ変換する方法 シーケンサーからのデータ転送 ランフォルダ構造と主要ファイル

RNA 6 - RNA MLP Back-propagation5

Benasque RNA 2012: RNA Motifs

Inhibition of RNA binding to hepatitis C virus RNA-dependent RNA

HEREDITARY TEST REQUISITION Fast Processing Prenatal ... · Blood-Adult(2 x 4ml EDTA) 4005 ACG ... charged as a solo. Reporting and data exchange .fastq .bam .vcf. Raw data (.fastq

RNA polymerase #1 General properties E. coli RNA polymerase Eukaryotic RNA polymerases

MSc-Modul RNA-Biochemie file•small noncoding RNA •Influenza –ein RNA-Virus RNA Biochemie 02/1 MSc-Modul RNA-Biochemie. Protein-Welt RNA-Welt Prä-RNA-Welt RNA Biochemie 02/2

Illumina Output Fastq format and Quality control · Fastq format – fasta with qualities • p = the probability that the corresponding base call is wrong • Qualities – p = 0.1

Transcription. Define the following Messenger RNA Ribosomal RNA Transfer RNA Transcription Codon

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RNA: Structure & Function. What is RNA? RNA is a nucleic acid called Ribonucleic Acid Functions of RNA: RNA transfers genetic information from the nucleus

Tipos de rna y rna ribosómico (r rna)

Transkriptiossa syntyy m-rna (lähetti-rna)

RNA-seq data analysis Project QI LIU. From reads to differential expression Raw Sequence Data FASTQ Files Unspliced Mapping BWA, Bowtie Mapped Reads SAM/BAM

Transcription: RNA synthesis, RNA processing, “Algorithms ... · Transcription: RNA synthesis, RNA processing, Intron-exon structure Promoters: Regulation, Chromatin structure,

RNA PROCESSING AND RNPs. RNA Processing Very few RNA molecules are transcribed directly into the final mature RNA. Most newly transcribed RNA molecules

Turbo-Decoding of RNA Secondary Structuregsharma/.../SharmaTurboDecodingRNASe… · RNA Structure Analysis: Motivation and Background RNA, noncoding RNA, RNA structure and its significance

RNA and Protein Synthesis · Web viewamino acid, anticodon, codon, gene, messenger RNA, nucleotide, ribosome, RNA, RNA polymerase, transcription, transfer RNA, translation Prior Knowledge

Eukaryotic RNA processing: alternative RNA splicing

RNA-Regulation: RNA Interference

1 NOTES: Chapter 13 - RNA & Protein Synthesis Vocabulary: Messenger RNA (mRNA) Ribosomal RNA (rRNA) Transfer RNA (tRNA) Transcription RNA Polymerase Codon

RNA Metabolism DNA-dependent synthesis of RNA RNA processing RNA-dependent synthesis of RNA & DNA

RNA Molecules and RNA Processing Functions and Modifications of RNA Molecules

SO YOU WANT TO DO A: RNA-SEQ EXPERIMENT Bioinformatics · 2016. 6. 17. · RNA-SEQ PIPELINE OVERVIEW Experiment Metadata Software Setup Sequence Data (fastq) preprocess Reference

skewer run2 SW041 1-trimir2.fastq FastQC Report · 2015. 5. 21. · Filename skewer_run2_SW041_1-trimmed-pair2.fastq File type Conventional base calls Encoding Sanger / Illumina 1.9

2011 RNA-RNA and RNA-protein interactions in coronavirus replication and transcription

RNA Matrices and RNA Secondary Structures€¦ · RNA Matrices and RNA Secondary Structures ... Applications: RNA in Biology, Bioengineering and Nanotechnology, University of Minnesota

Le contrôle qualité sur les données fastq