Skip to content
Permalink
290e9c18f6
Switch branches/tags

Name already in use

A tag already exists with the provided branch name. Many Git commands accept both tag and branch names, so creating this branch may cause unexpected behavior. Are you sure you want to create this branch?
Go to file
 
 
Cannot retrieve contributors at this time

RNA-Seq pipeline

Converting Illumina run to FASTQ

bcl2fastq 
	--runfolder-dir <path-to-nextseq-run>
	--output-dir <path-to-save-fastq-samples>
	--no-lane-splitting
	--loading-threads 8
	--demultiplexing-threads 8
	--writing-threads 8
	--sample-sheet /path/to/SampleSheet.csv

Aligning reads to genome

START
	--runMode alignReads
	--runThreadN 8
	--genoeDir <path-to-genome-star-index>
	--readFilesIn <path-to-fastq-file>
	--readFilesCommand zcat
	--outStd Log
	--outSAMtype BAM SortedByCoordinate
	--outSAMstrandField intronMotif
	--outFilterIntronMotifs RemoveNoncanonical
	--alignSoftClipAtReferenceEnds No
	--outFilterScoreMinOverLread 0.25
	--outFilterMatchNminOverLread 0.25

Note: sometimes depend on your seq. library you will need to update the parameters to optimise the alignment. More parameters can be found in the manual.

Annotate alignments

featureCounts
	-a <path-to-GTF-genome-annotation>
	-o <path-to-save-counts>/counts.txt
	-t exon
	-Q 255
	-T 8
	$(ls <path-to-BAM-file>/*.bam)

Differential expressions with R

Other useful tools