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== Alignment with Bowtie2 == * The long reads were not compatible with our pipeline using Bowtie2, because the aligner suppresses read name lines which are longer than 256 characters. This causes truncated original reads stored in the read name line. * Added code to split long reads into multiple short reads to use Bowtie2 * [[File:smartBisReadMapper.txt]] * Mapping pipeline: <nowiki> cur_dir="/media/3TB_Dinh/Blueprint" reads_dir="/media/3TB_Dinh/Blueprint" email="diep.hue.dinh@gmail.com" bisReadMapper="/home/ddiep/scripts/MethylationPipeline/scripts/smartBisReadMapper.pl" template_fwd="/media/2TB_storeA/BisRef/bisHg19/hg19.fa.bis.fwd.bowtie2" template_rev="/media/2TB_storeA/BisRef/bisHg19/hg19.fa.bis.rev.bowtie2" template_fa="/media/2TB_storeA/BisRef/bisHg19/hg19.fa" soap="/home/ddiep/softwares/soap2.21release/soap" bowtie="bowtie2" cpg="/media/2TB_storeA/BisRef/bisHg19/C_Pos/hg19.fa.cpg.positions.txt" INDX="1Index1_S1 1Index2_S2 1Index3_S3 2Index4_S6 1Index5_S4 2Index6_S7 2Index7_S8 1Index8_S5" # make sure the qual_base variable is set correctly cd $cur_dir for s in ${INDX} do f="$s.fq" g="$s.leftR1.fq" h="$s.leftR2.fq" n="$s-MS" mkdir $n echo "cd $cur_dir/$n" > $n.job echo "$bisReadMapper -r $reads_dir/$f -W $template_fwd -C $template_rev -g $template_fa -a $bowtie -p 16 -b 33 -n $n.f1 -q 20 -l $cpg > $n.f1.statusMbias 2>$n.f1.err" >> $n.job echo "$bisReadMapper -r $reads_dir/$g -W $template_fwd -C $template_rev -g $template_fa -a $bowtie -p 16 -b 33 -n $n.f2 -q 20 -l $cpg > $n.f2.statusMbias 2>$n.f2.err" >> $n.job echo "$bisReadMapper -r $reads_dir/$h -W $template_fwd -C $template_rev -g $template_fa -a $bowtie -p 16 -b 33 -n $n.f3 -q 20 -l $cpg > $n.f3.statusMbias 2>$n.f3.err" >> $n.job echo "rm *encoded" >> $n.job done </nowiki> ===Mapping statistics=== * f1 = COPE combined PE reads, f2 = leftover R1, f3 = leftover R2. * With clipping, we got 2.846 Gbps mapped, without clipping, we got 2.842 Gbps mapped. Clipping did not improve the amount of usable bp by a lot. * Libraries Index 7 and Index 8 have the best mapping rates. {| {{table}} | align="center" style="background:#f0f0f0;"|'''index''' | align="center" style="background:#f0f0f0;"|'''read file''' | align="center" style="background:#f0f0f0;"|'''total clipped bases''' | align="center" style="background:#f0f0f0;"|'''total clipped mapped bases''' | align="center" style="background:#f0f0f0;"|'''%mapped(clipped)''' | align="center" style="background:#f0f0f0;"|'''total bases''' | align="center" style="background:#f0f0f0;"|'''total mapped bases''' | align="center" style="background:#f0f0f0;"|'''%mapped''' |- | 1||f1||8255745||5487013||66.46%||16078514||9619049||59.83% |- | 1||f2||79964964||53864094||67.36%||104318024||50945748||48.84% |- | 1||f3||89749717||50800000||56.60%||99998101||48797421||48.80% |- | 2||f1||16566696||11031637||66.59%||32606092||19771959||60.64% |- | 2||f2||171105395||112357034||65.67%||230586489||106229965||46.07% |- | 2||f3||189062773||102110571||54.01%||215612917||97902718||45.41% |- | 3||f1||25104735||15365281||61.20%||51734457||30631757||59.21% |- | 3||f2||285378749||171910645||60.24%||405686018||161655992||39.85% |- | 3||f3||330732139||151852920||45.91%||372674273||145065779||38.93% |- | 4||f1||21403635||13378820||62.51%||42833085||24714027||57.70% |- | 4||f2||222330667||138200084||62.16%||300568658||131048023||43.60% |- | 4||f3||250430069||125847956||50.25%||281078201||120620387||42.91% |- | 5||f1||38452894||26019949||67.67%||67355222||42886852||63.67% |- | 5||f2||360645974||247314571||68.58%||463290839||235545215||50.84% |- | 5||f3||401191540||226901252||56.56%||438788886||218980079||49.91% |- | 6||f1||30067978||18916755||62.91%||56145035||34232803||60.97% |- | 6||f2||289462431||191584810||66.19%||389224257||183501240||47.15% |- | 6||f3||328112598||176045539||53.65%||364481484||169262473||46.44% |- | 7||f1||32538699||22178956||68.16%||49348430||34280682||69.47% |- | 7||f2||253380410||192127161||75.83%||313644714||186792389||59.56% |- | 7||f3||279659004||180487623||64.54%||300856044||175441163||58.31% |- | 8||f1||50524286||33292157||65.89%||81821672||53601105||65.51% |- | 8||f2||415435035||301628773||72.61%||516798645||291900532||56.48% |- | 8||f3||463261828||277764207||59.96%||496294120||269456511||54.29% |}
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