Help with ERROR: Paired-end reads were detected in single-end read library

Question

[Chris]

Hi all, I run a pretty similar command for two RNA-seq data sets:

featureCounts -a Homo_sapiens.GRCh38.107.gtf -o count.out -T8 /mapped/*.bam

I only change the bam files and both RNA-seq data are paired-end reads. However, one command got the error above and the other is not. Would you please have a suggestion? Thank you so much!

Answer 1

[Mensur Dlakic]

We know that a program is operating as intended, and that .gtf file is good. A simple deduction is that something is wrong with your .bam files from the second group. It is difficult to tell what is wrong from where we stand because we don't know the error message, and also because we don't know about the differences in how the .bam files were created. An educated guess is that the mapping was done against a different reference genome.

[Chris]

Thank you for your reply. My star command used the same --genomeDir that I checked are the same. Is that because of the data? The command I used to get the bam file:

STAR --runThreadN 32 --runMode alignReads --genomeDir /genomeDir --readFilesIn /1.fq /2.fq --outSAMtype BAM Unsorted --outFileNamePrefix aligned