differences between featureCounts and htseq-count

Question

[golany89]

Hi, I have RNA-seq data (from ribodepleted samples) and i'm trying to map my reads to features using either htseq or featureCounts. The input for both is a bam file (reads were aligned using STAR) and a GTF file from ENSEMBLE. Both assign reads to exons by default.

FeatureCounts reported that ~33% of my reads were assigned to features, however when I'm trying to check how many reads were assigned using htseq, I saw that only 2% of my reads were assigned (There is no report in htseq so I summerized the second column in the results file, without the last 5 rows that contain the "no-feature" and "ambiguous" etc).

What is causing this difference between the two algorithms? Thank you very much!

[igor]

The differences should be very minor between the two tools. If you post the exact commands used, it will be easier to see what causes the differences.

[colin.kern]

Is it possible you're using the wrong stranded argument in htseq? The vast majority of stranded libraries (e.g. using the Illumina kit) should use --stranded=reverse NOT --stranded=yes

[golany89]

Thank you very much, this solved the problem.

Answer 1

[Arup Ghosh]

Comparison Htseq And Feature Count