[tsalem]

When you did QC/trimming of the data was a PE-read aware program used (to keep the pairs of reads in sync in both files) with the two data files at the same time (R1/R2)? It is possible that reads in your files went out of sync and this is resulting in the error above.

Actually, I didn't do any thing with the files, I just got the .gtf file from The ucsc.
and the .bam file from my advisor. and i am trying to get genes count from .bam file.

is possible to let me know how i can check if the pairs of reads in sync in both files

Thanks

[GenoMax]

Do you know how the bam was generated (is there more than 1 bam per sample)? Do you have access to the original read files?

[tsalem]

I don't know how it has been generated, i have different samples and there is one bam file for every sample.

I don't have access to the original read files.

[tsalem]

I got the solution I was using Wrong .gtf file.

Thanks Geno for looking at this.