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Kun:LabNotes/SingleCellExpr/2011-10-6
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==Repeat analysis of the 10 PD brain RNAseq data== ===Tophat mapping=== nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 1741_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 1741_cDNA_sequence.txt > 1741.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 1818_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 1818_cDNA_sequence.txt > 1818.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 1947_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 1947_cDNA_sequence.txt > 1947.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 4526_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 4526_cDNA_sequence.txt > 4526.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 4735_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 4735_cDNA_sequence.txt > 4735.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 4789_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 4789_cDNA_sequence.txt > 4789.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 4879_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 4879_cDNA_sequence.txt > 4879.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 4977_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 4977_cDNA_sequence.txt > 4977.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 5028_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 5028_cDNA_sequence.txt > 5028.log& nohup /home/kunzhang/softwares/tophat-latest/tophat -p 4 --solexa1.3-quals -o 5171_tophat -G /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf /GenomeDB/UCSC/Homo_sapiens/hg19/Sequence/BowtieIndex/genome 5171_cDNA_sequence.txt > 5171.log& ===Cuffdiff analysis=== /home/kunzhang/softwares/cufflinks-1.0.3.Linux_x86_64/cuffdiff /GenomeDB/UCSC/Homo_sapiens/hg19/Annotation/Genes/genes.gtf \\ -N -p 8 -o PD_vs_control 1741_tophat/accepted_hits.bam,1947_tophat/accepted_hits.bam,4526_tophat/accepted_hits.bam,4879_tophat/accepted_hits.bam,4977_tophat/accepted_hits.bam \\ 5028_tophat/accepted_hits.bam,5171_tophat/accepted_hits.bam,1818_tophat/accepted_hits.bam,4735_tophat/accepted_hits.bam,4789_tophat/accepted_hits.bam *In the gene_exp.diff file, I selected the genes that were significantly different between PD and the controls, and filtered out those that either didn't express or expressed extremely low (FPKM<0.01) in one group. I ended up getting 38 genes that were up-regulated in the controls, and 37 that were down-regulated in the controls. The majority of these genes turned out to be microRNA genes. I'm not sure how to interpret this observation since microRNAs were not supposed to be captured in this RNAseq protocol.
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