I'm a beginner in RNAsequencing analysis, so I apologize for naive questions :)
I have single end RNAseq datas from human prepared by NEBNext® UltraTM II Directional RNA Library Prep Kit for Illumina. I used STAR for alignments and now I would like to summary readCount using FeatureCounts.
So I would like to use:
featureCounts \ -t exon -g gene_id \ -s ??????? \ -a ./Annotation/Homo_sapiens.GRCh38.84.chr.gtf \ -o ./Trim/Sample1_Count_Gene.txt \ ./Trim/Sample1_Aligned.sortedByCoord.out.bam
My problem come from the -s argument in FeatureCount which need the strand specificity of the read. I try the 3 possibilities (0,1,2) and 0 and 2 works well but 1 option give me low assigned read.
I'm not sure but,I think that if we use dUTP, it maintains the reverse strand of amplified cDNA so it means that I have to use -s2 in my analysis? Sorry maybe I'm totally wrong ...