第23期培訓課件雙擊桌面上的圖標putty exe后顯示窗口

“HostName:”IP192.168.118.204”[lirj@big[lirj@big~]$pwd 當前位置更換到我們事先已經(jīng)為您建好的計算下面。我們在光標所在的位置鍵入（例如：你的用戶名是lirj：[lirj@big[lirj@big~]$ BSMAP軟件的地址： 文章在BMCBioinformatics2009, BSMAP[lirj@big[lirj@biglirj]$ls 接下來關于BSMAP軟件的運算練習將在這個 ./[lirj@bigBSMAP]$ls readlengthuptoallowupto20mismatchessupportpairendmapsupportparallelmapqueryfilebforpairenddata,FASTA/FASTQ/BAMformat.IftheinputisinBAMformat,itshouldbethesameasthefilespecifiedby"-a"outputalignmentfile,iffilenamehas.samsuffix,theoutputwillbeinSAMformat,ifthefilenamehas.bamsuffix,theoutputfilebeinsortedBAMfile,andafilename.baiindexfilewillbegenerated,forotherfilenamesuffixtheoutputisinBSPoutputalignmentfileforunpairedreadsinpairendmap,onlyusedforBSPformatoutput.IftheoutputformatisspecifiedinBAM/SAMformat,thisoptionwillbeignored,allalignmentswillbewritentooneBAM/SAMoutputfilespecifiedbythe"-o"seedsize,default=16,min=8,[seed_size*seed_seg_num+3<=read_length]longerseedsizeisfaster,~1.5timesfasterwitheachadditionalntmaxnumberofmismatchesallowedonaread,default=2,maxmaxnumberofequalbesthitstocount,smallerwillbeinitialquality,[Illuminauses'@',SangerInstituteuses'!'],filterlow-qualityreadscontaining>nNs,numberofprocessorstouse,maxinsertionsizeforpairendmap,mininsertionsizeforpairendmap,indexinterval(1~16),default=4,meaningthereferencegenomewillbeevery4bp,largerindexintervalneedslessmemory,andslightlymapsensitivity.(~0.5%3-endadaptersequence,default=none,requiresatleast4ntmatched,mismatchincludethereferencesequencesastheXR:Z:<string>fieldinSAMdefault=donotstartfromthenthreadorreadpair,default:endatthenthreadorreadpair,default:setrestrictionenzymedigestionsiteandactivateRRBSmapmode,readsmustbemappedtodigestionsitesthedigestionsitemustbepalindromic,digestionpositionismarkedby'-',forexample:'-DC-CGG'(MspI)randomeseedinselectingmultiplehits.default:0(seedsetfromsystemsingleend在 [lirj@big[lirj@bigBSMAP]$bsmap-areads1.fastq-dchr22.fa-o-w100-s14-v3[lirj@big[lirj@bigBSMAP]$psPIDPIDTTYTIME3399pts/300:00:007820pts/300:00:037826pts/300:00:0010000readsfinished.5secs10000readsfinished.5secs20000readsfinished.5secsTotalnumberofalignedreads:20000(1e+02%)FinishedatThuSep Totaltimeconsumed: 5secs bsmap-areads1.fastq-dchr22.fa-o_bsmap.bsp-w100-14-v[lirj@big[lirj@bigBSMAP]$ls可以看到，文件夾里多了一個文件map_bsmap.bsp，這就是運行BSMAP將[lirj@big[lirj@bigBSMAP]$lessS _bsmap.bsp屏幕上將顯示輸出文件的內(nèi)容1read2mappedread3qualityofthequery4UM:uniquemapOF:overmapNM:nomapQC:lowquality56maplocation(1based,5'-endcoordinatesoftheregionontheWatsonstrandof7++:forwardstrandofWatsonofreference+-:reversestrandofWatsonofreference-+:forwardstrandofCrickofreference--:reversestrandofCrickofreference8insertionsizeforpair-endmap,0meanssingle-endunpairedmap9Wastonreferencesequenceatthemapnumberofmismatchesofcurrent#hitsof0mismatchto#hitsofmax_mismatches,by在光標處鍵入“q”，回到 在 [lirj@big[lirj@bigBSMAP]$bsmap-areads1.fastq-dchr22.fa-o-w100-s14-v3BSMAPBSMAPStart Thu 118:42:29Loadin1dbseqs,totalsize bp.2secspassedCreateseedtable.5secsmaxmismatches:startfromread#1maxmulti-hits:maxNs:seedsize: basequalitychar:maxfragemtQuery:reads1.fastqReference:chr22.faOutput:map 0000readsfinished.5secspassed20000readsfinished.5secsFinishedatThu 118:42:34 5 bsmap-areads1.fastq-dchr22.fa-o_bsmap.sam-w100-14-v[lirj@big[lirj@bigBSMAP]$ls可以看到，文件夾里多了一個文件map_bsmap.sam，這就是運行BSMAP將[lirj@bigBSMAP]$[lirj@bigBSMAP]$lessS _bsmap.sam1QuerypairNAMEifpaired;orQueryNAMEif2UM:0x0,MA:0x100,OF:0x100,NM:0x4,QC:formaponBSCorBSWC:FLAG=FLAG+0x10forpair-endmap:ifit'sthefirstreadinpair,ifit'sthesecondreadinpair,FLAG=FLAG+0x80ifmapsarepaired,FLAG=FLAG+0x2ifmateisunmapped,ifmateismappedonBSCorBSWC,3Referencesequence41-basedleftmostPOSition/coordinateoftheclipped5MAP6extendedCIGAR7MateReferencesequenceNaMe;“=”ifthesameas81-basedleftmostMatePOSitionoftheclipped9inferredInsertqueryqueryXR:Z:<referencesequence>從map位置開始的waston參考序列在 [lirj@big[lirj@bigBSMAP]$bsmap-areads1.fastq-dchr22.fa-o-w100-s14-v3BSMAPBSMAPStart Thu 118:44:15Loadin1dbseqs,totalCreateseedtable.4secsbp.1secsmaxmismatches:startfromread#1maxmulti-hits:maxNs:seedsize: basequalitychar:maxfragemtQuery:reads1.fastqReference:chr22.faOutput:map 10000readsfinished.4secspassed20000readsfinished.4secsFinishedatThuSep Totaltimeconsumed:4secsConvertingSAMtoBAM...[samopen]SAMheaderispresent:1sequences.SortingBAM...IndexingBAM bsmap-areads1.fastq-dchr22.fa-o_bsmap.bam-w100-14-v[lirj@big[lirj@bigBSMAP]$ls這就是運行BSMAP將重亞硫酸鹽處理后的短序列文件比對到組后的[lirj@big[lirj@bigBSMAP]$samtoolsview _bsmap.bam|lessS可以節(jié)約空間，可用samtools工具查看。pairend在 [lirj@big[lirj@bigBSMAP]$bsmap-areads1.fastq–breads2.fastq-dchr22.fa-_bsmap_pairend.bsp-2_bsmap_unpair.bsp-w100-s14-v3single180single180bsmap-areads1.fastq-breads2.fastq-dchr22.fa-_bsmap_pairend.bsp-2_bsmap_unpair.bsp-w100-s14-v[lirj@big[lirj@bigBSMAP]$lsmap_bsmap_unpair.bsp，這就是運行BSMAP將重亞硫酸鹽處理后的pairend短序列文件比對到組后的生成文件。那么我們現(xiàn)在看看比對結(jié)果，我們 _bsmap_pairend.bsp屏幕上將顯示輸出文件的內(nèi)容在 [lirj@big[lirj@bigBSMAP]$bsmap-areads1.fastq–breads2.fastq-dchr22.fa-o _bsmap_pairend.sam-w100-s14-v3&single180single180bsmap-areads1.fastq-breads2.fastq-dchr22.fa-_bsmap_pairend.sam-w100-s14-v[lirj@big[lirj@bigBSMAP]$lsBSMAP將重亞硫酸鹽處理后的pairend短序列文件比對到組后的生成[lirj@big[lirj@bigBSMAP]$lessS _bsmap_pairend.sam屏幕上將顯示輸出文件的內(nèi)容在 [lirj@big[lirj@bigBSMAP]$bsmap-areads1.fastq–breads2.fastq-dchr22.fa-o _bsmap_pairend.bam-w100-s14-v3&single180single180bsmap-areads1.fastq-breads2.fastq-dchr22.fa-_bsmap_pairend.bam-w100-s14-v[lirj@big[lirj@bigBSMAP]$ls[lirj@big[lirj@bigBSMAP]$samtoolsview _bsmap_pairend.bam|lessS屏幕上將顯示輸出文件的內(nèi)容RMAP軟件的地址：文章在 Bioinformatics2009,25:2841 接下來我們練下 到BSMAP的上一級 RMAP[lirj@biglirj]$ls[lirj@biglirj]$ls RMAP ./[lirj@bigRMAP]$ls就會看到這一 下面有三個文件，分別是chr22.fa，reads1.fastq -o,-Nameofoutputfile(default:-c,-FASTAfileordircontaining-S,-numberofwidthof-m,-umallowed-M,-max-umallowedmapsfora-Q,-usequalityscores(inputmustbeprintmorerun-f,-fasterseeds(sensitiveto2[lirj@big[lirj@bigRMAP]$reads1.fastq& -c 12在此上的起始位34reads56比對到了參考序列的哪一股（正或負[lirj@big[lirj@bigRMAP]$ -c mapped_bs_locations2.bedBSSeeker軟件的地址：Seeker/BS文章 BMCBioinformatics2010,接下來我們練下BSSeeker的使用方法。目前我們還在RMAP [lirj@bigRMAP]$[lirj@bigRMAP]$cd..到RMAP的上一級 [lirj@big[lirj@biglirj]$ BS_Seeker[lirj@biglirj]$ls [lirj@big[lirj@biglirj]$ / ./命令：pythonPreprocessing_genome.pyshowthishelpmessageandInputyourreferencegenomefilename(fastaReadscontainingtags? PathtoBowtie[~/bowtie-Add">log_Preprocessing_genome.txt"attheendofcommandstosavetheIDassignmentstothereferencesequences[lirj@big[lirj@bigBS_Seeker]$ chr22.fa log_Preprocessing_genome.txt&1465900:00:001821200:00:021821500:00:00 Done-f [lirj@big[lirj@bigBS_Seeker]$ls窗口顯示 reference_genome第二步，比對組-h,--showthishelpmessageandInputyourreadfile(supportinformat:Solexaseq,sequences,illuminafastq,qseq)Readscontainingtags?Y/N ifreadshavetags:-fFWtag-rRCtagThelastcyclenumberofyourreadtobemappedPathtoBowtie[~/bowtie-PathtoReferencegenomelibraryNumberofmismatches(0,1,2,3)[lirj@big[lirj@bigBS_Seeker]$ - N-e36- -o[lirj@big[lirj@bigBS_Seeker]$ls我們會發(fā)現(xiàn)，我們多了兩個文件：log_myoutput.txt myoutput.txt1ReadID(fromthefirst4columnsinSolexaseqfile,oraserialnumberoftheoriginalinput)2NumberofmismatchesbetweenthegenomicseqandtheBSreadlistincolumns6and7.ThebisulfiteconvertedsitesbetweenreadTstogenomicCsarenotincluded.3Thestrandwhichthereadmaybefrom(+FW,+RC,-RC,-4Thecoordinateofthemappedposition:thefirst4digits(ID)indicatethechromosome,the"+"or"-"indicatethemappedstrand.Thelast10digitsarethe1-based,5'-endcoordinateofthemappedgenomicsequenceontheWatsonstrand.5Thegenomicsequenceofthemappedregionplus+2and-26BSreadsequencesfrom5'to3':ifthereadsareuniquelymappedastheywereFWreads,theoriginalreadsareshown.IfthereadsareuniquelymappedastheywereRCreads,theirre