HTseq-count error: mate pairs cannot be matched

[paulb5677]

Hi guys, I'm having some trouble running HTseq-count on paired end RNA-seq data. I use the following command to run HTse-count:

htseq-count -s no -a 10 Sorted.sam Homo_sapiens.GRCh37.74.gtf > Sorted.count

And the following command is used to sort the bam prior to running HTseq-count:

samtools sort -n bamfile.bam sorted_bamfile

There seems to be a problem with matching paired reads. The following error message is reported and empty .count files are generated:

Warning: Read HISEQ:309:C4UR9ACXX:2:1101:2467:41039 claims to have an aligned mate which could not be found. (Is the SAM file properly sorted?)

And the relevent section of the SAM file looks like this:

HISEQ:309:C4UR9ACXX:2:1101:2467:41039	81	X	71494941	3	50M	19	22189602	0	GACCCTTGGTGTCATAGATCAGACGGAAATTCTCTCCCGTCTTGTCAATG	FBIIIIIIIIIIIIIIIIIIIIIIIIIIIIFFFIIIIFFFFFFFFFFBBB	AS:i:0	XN:i:0	XM:i:0	XO:i:0	XG:i:0	NM:i:0	MD:Z:50	YT:Z:UU	XS:A:-	NH:i:2	CC:Z:=	CP:i:71494941	HI:i:0
HISEQ:309:C4UR9ACXX:2:1101:2467:41039	337	X	71494941	3	50M	19	22394287	0	GACCCTTGGTGTCATAGATCAGACGGAAATTCTCTCCCGTCTTGTCAATG	FBIIIIIIIIIIIIIIIIIIIIIIIIIIIIFFFIIIIFFFFFFFFFFBBB	AS:i:0	XN:i:0	XM:i:0	XO:i:0	XG:i:0	NM:i:0	MD:Z:50	YT:Z:UU	XS:A:-	NH:i:2	HI:i:1

Has anyone else experienced this? Do you think its a problem with the fastq files, Tophat, Samtools, or HTseq-count?

Thanks