轉(zhuǎn)錄組分析流程salmon瑞客

(d)不基于比對，

&易o(hù)m生信e，b畢io生lo緣g；y.培b訓(xùn)io版m權(quán)e所d有c xx

(.fastq

(.fafile)

(.gtffile)(可選)

A FASTQxx

CDNA (.fa (.gtf (.fastq

FASTQqualityscores=ASCIIqualityscore-either33or Illumina e=estimatedprobabilityofthebasecallbeingwrongQ40:1errorin10,000basecalls;Q30:1errorin1,000;Q28:1.6errorin1000;Q25:3errorin1000;Q20:1errorin信，畢生緣；培訓(xùn) FASTQ CDNA (.fa (.gtf xx 信，畢生緣；培訓(xùn) 公共數(shù)據(jù)從NCBISRA使用NCBI提供的SRA-toolkit中的工具fastq-dump直接SRR文件，并轉(zhuǎn)換雙端就自動(dòng)拆分，如果是單端不受的數(shù)據(jù)集一般比較大，放入 (nohupcmd&)。nohupfastq-dump-v--split-3--nohup&fastq-dump-v--split-3--& 77 fastqc x-axis:Positionin y-axis:Quality信，畢生緣；培訓(xùn) (箱線圖通過最大值、上四分位示質(zhì)量得分。將所有reads的第一位堿基質(zhì)量得分進(jìn)行箱線 信，畢生緣；培訓(xùn) 信，畢生緣；培訓(xùn) 每個(gè)readGC含量的分布圖。橫坐標(biāo)表示平均GC含量，縱坐標(biāo)表示reads數(shù)。左圖顯示每個(gè)read的GC分布(紅線)與理論分布(藍(lán)線)相契合，GC含量均一。右圖出現(xiàn)了GC含量雙峰，表示樣品可能存在特定的序列污染如混入了引物二聚體，當(dāng)這一信，畢生緣；培訓(xùn) ../FastQC/Configuration/adapter_list.txtFastQC工具會與這兩個(gè)文件里的序列進(jìn)行比信，畢生緣；培訓(xùn) 信，畢生緣；培訓(xùn) 直方圖展示有 量異常高或低的樣本 橫軸是GC含量的判斷，縱軸是堿基橫軸表 reads數(shù)，縱軸表示給定 數(shù)區(qū)間內(nèi)樣本數(shù)目，即給 深度有多 樣本 reads數(shù)低于20M的樣品沒有， 高于40M

信，畢生緣；培訓(xùn) 接頭和低質(zhì)量readsmCommondjava-jartrimmomatic-0.30.jarPE--phred33input_forward.fqinput_reverse.fqoutput_forward_paired.fqoutput_forward_unpaired.fqoutput_reverse_paired.fqoutput_reverse_unpaired.fqILLUMINACLIP:adaptor-PE.fa:2:30:10LEADING:20TRAILING:20MINLEN:36TwoinputfilesandfouroutputRemoveadapters:( um2mismatchesinthe'seed'(16bases)oftheadaptor;palindromeclipthreshold30;simpleclipthreshold10)Removeleadinglowqualitybases(belowquality20);Removetrailinglowqualitybases(belowquality20);Dropreadsbelowthe36base易s生信lo，n畢 (2x(2x150(.fastq

(.fafile)

(.gtf 信，畢生緣；培訓(xùn) Ensembl(信，畢生緣；培訓(xùn)E。nsemblcDNA指編 的 RNA- GTF(GeneTransferFormat, RNA- CDNA(.faCDNA(.fa(2x150(.fastq 易s生c信.，e畢d生u緣/F；A培Q訓(xùn)版/F權(quán)A所Q有 m1-seqname-nameofthechromosomeorscaffold;chromosomenamescanbegivenwithorwithoutthe'chr'prefix.Importantnote:theseqnamemustbeoneusedwithinEnsembl,i.e.astandardchromosomenameoranEnsemblidentifiersuchasascaffoldID,withoutanyadditionalcontentsuchasspeciesorassembly.SeetheexampleGFFoutputbelow.m2-source-nameoftheprogramthatgeneratedthisfeature,orthedatasource(databaseorprojectname)m3-feature-featuretypename,e.g.Gene,Variation, 4-start-Startpositionofthefeature,withsequencenumberingstartingat1. 5-end-Endpositionofthefeature,withsequencenumberingstartingat1. 6-score-Afloatingpointvalue. 7-strand-definedas+(forward)or- 8-frame-Oneof'0','1'or'2'.'0'indicatesthatthefirstbaseofthefeatureisthefirstbaseofacodon,'1'thatthesecondbaseisthefirstbaseofacodon,andsoon.. 9-attribute-Asemicolon-separatedlistoftag-valuepairs,providingadditional6612信，畢生緣；培訓(xùn) bed–至少3列，另外9列可選，0-start信，畢生緣；培訓(xùn) 信，畢生緣；培訓(xùn)

- (.gtffile)(.gtffile)(.fafile)(.fastqx RNA- A transcripts_index--typequasi-k <LIBTYPE>-1reads1.fq-2reads2.fqo2 定量時(shí)考慮到不同樣品 長度的改變（比如不同 g易n生e信t，te畢s生/緣D；E培訓(xùn)S版e權(quán)q所2有/。 (.fafile)(.fastqx(.fafile)(.fastqx RNA- (.gtf 信，畢生緣；培訓(xùn) (.fafile)(.fafile)(.fas