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==Checking RNA-Seq Read Coverage of Probe Targets== *[[Matt:LabNotes/2014-11-1#Filter_Probes_with_Low_Coverage_based_on_Brain_RNA-seq_Data_from_Rui.2FBlue | Probes were already filtered for those with low coverage in bulk tissue RNA-Seq with all Brodmann Areas summed together]] **Still, we can check just BA8 to see if that explains lack of detection of certain genes ===Create BED File for Probe Targets=== *[[Media:CA12kNov2014_Probelist2BED.txt |CA12kNov2014_Probelist2BED.pl]] **Made start and end positions +/- 25bp from target position **[[Media:OutputFile_0gap_contig_final.full_info.V4.txt|OutputFile_0gap_contig_final.full_info.V4.txt]] > [[Media:CA12k_Nov2014_V4.txt|CA12k_Nov2014_V4.bed]] **[[Media:OutputFile_0gap_final.full_info.V7.txt|OutputFile_0gap_final.full_info.V7.txt]] > CA12k_Nov2014_V7.bed *Convert from hg38 reference positions to hg19 reference (which RNA-Seq data was mapped to) using [http://genome.ucsc.edu/cgi-bin/hgLiftOver UCSC LiftOver] **CA12k_Nov2014_V4.bed > CA12k_Nov2014_V4_hg19.bed **CA12k_Nov2014_V7.bed > CA12k_Nov2014_V7_hg19.bed ===Bedtools Coverage with RNA-Seq Data=== *Workspace **genome-miner:/home/mzcai/CA12kNov2014_V4_CaptureAnalysis **genome-miner:/home/mzcai/CA12kNov2014_V7_CaptureAnalysis *RNA-Seq data from: /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR$ **RL-BA8-sec9-t-N701-15May14_S1_mapped/RL-BA8-sec9-t-N701-15May14_S1_Aligned.sorted.bam ***59,038,666 reads **RL-BA8-sec9-n-N702-15May14_S2_mapped/RL-BA8-sec9-n-N702-15May14_S2_Aligned.sorted.bam ***45,222,268 reads *Commands <!-- /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -hist -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-t-N701-15May14_S1_mapped/RL-BA8-sec9-t-N701-15May14_S1_Aligned.sorted.bam -b CA12k_Nov2014_V4_hg19.bed > coverage.hist.BA8-t.V4.txt & /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -hist -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-n-N702-15May14_S2_mapped/RL-BA8-sec9-n-N702-15May14_S2_Aligned.sorted.bam -b CA12k_Nov2014_V4_hg19.bed > coverage.hist.BA8-n.V4.txt & /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -counts -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-t-N701-15May14_S1_mapped/RL-BA8-sec9-t-N701-15May14_S1_Aligned.sorted.bam -b CA12k_Nov2014_V4_hg19.bed > coverage.counts.BA8-t.V4.txt & /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -counts -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-n-N702-15May14_S2_mapped/RL-BA8-sec9-n-N702-15May14_S2_Aligned.sorted.bam -b CA12k_Nov2014_V4_hg19.bed > coverage.counts.BA8-n.V4.txt & --> /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-t-N701-15May14_S1_mapped/RL-BA8-sec9-t-N701-15May14_S1_Aligned.sorted.bam -b CA12k_Nov2014_V4_hg19.bed > coverage.BA8-t.V4.txt & /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-n-N702-15May14_S2_mapped/RL-BA8-sec9-n-N702-15May14_S2_Aligned.sorted.bam -b CA12k_Nov2014_V4_hg19.bed > coverage.BA8-n.V4.txt & /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -counts -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-t-N701-15May14_S1_mapped/RL-BA8-sec9-t-N701-15May14_S1_Aligned.sorted.bam -b CA12k_Nov2014_V7_hg19.bed > coverage.BA8-t.V7.txt & /home/mzcai/softwares/bedtools-2.20.1/bin/coverageBed -counts -abam /media/LTS_33T/RL_LTS33T/201404_201405_7Samples_BulkNucleiBatch1-20140623_Expt146/STAR/RL-BA8-sec9-n-N702-15May14_S2_mapped/RL-BA8-sec9-n-N702-15May14_S2_Aligned.sorted.bam -b CA12k_Nov2014_V7_hg19.bed > coverage.BA8-n.V7.txt & *Sum up counts for each gene: CollapseSum_BedtoolsCoverageCounts.pl **e.g. coverage.BA8-n.V7.txt > coverage.BA8-n.V7.collapsed.txt ===Results=== [[Media:20150427_BulkSeqCounts_CA12kNov2014Targets.xlsx]]
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