1. RSEM的安装和使用:
$ tar -xzf RSEM-1.3.0.tar.gz
$ cd RSEM-1.3.0
$ make
$ make ebseq
$ echo 'PATH=$PATH:/.../' >> ~/.bashrc
$ source ~/.bashrc
$ extract-transcript-to-gene-map-from-trinity trinity.fa gene_map (可选,转录本和基因都做)
$ rsem-prepare-reference --bowtie2 ref_trinity.fasta ref_trinity (I. Preparing Reference Sequences)
$ rsem-prepare-reference --transcript-to-gene-map ene_map --bowtie2 ref_trinity.fasta ref_trinity(可选,转录本和基因都做)
$ rsem-calculate-expression -p 48 --paired-end --bowtie2 read1.fq read2.fq ref_trinity sample.name (II. Calculating Expression Values)
$ rsem-plot-model sample.name sample.name.diagnostic.pdf (可选,看一些统计)
2. 差异表达:
$ rsem-generate-data-matrix sampleA.results sampleB.results ... > samples.matrix
$ rsem-run-ebseq samples.matrix 3,3 dge.result (两个组,每个组3个重复,如2,,3,3为3个组分别有2/3/3个重复)
$ rsem-control-fdr dge.result 0.05 dge.filter.result (controlling the false discovery rate (FDR) at level 0.05) (PostFC is recommended over the RealFC)