RNAseq analysis workflow (Under development)

Author: Priya Lakra

Quality control

Start with quality checking of the RNAseq fastq files. Commonly used tools include FastQC MultiQC

bash scripts/pl_qc.sh [options]

Trimming adaptors

QC may or may not be followed by adaptor trimming. Commonly used tool is trimmomatic

bash scripts/pl_trim.sh [options]

QC should also be performed after trimming the sequences!

Alignment

Multiple alignment tools are there in this world of bioinformatics. We will use here star.

bash scripts/pl_STARindexgen.sh [options]

bash scripts/pl_STARalign.sh [options]

Alignment step is tricky. Always read carefully the documentation before setting up any advanced paramaeters. This step can drastically affect downstream analyses. Alignments can be quality check using RSeQC, QoRTs, etc.

Read Count

Cool, we are done with alignment. We will now move to counting those alignments in a meaningful way. Few tools for counting reads are featurecounts and [HTSeq Count]....

I'm using here featureCounts

bash scripts/pl_counts.sh [options]

Differential gene expression analysis

Now we have the read count matrix. With this we can perform various downstream analyses such as Differential gene expression analysis, etc.

Using R packages

DESeq.R