Problems with htseq-count reading bam file created by STAR

[priya]

Hi,

Based on the STAR aligned bam files, I tried to generate the counts of RNA-seq data using htseq-count (version 0.5.1) tool by giving following command

htseq-count -m intersection-nonempty input_bam_file gtf_file > Sample1_count.txt

I am getting the following error

Error occured when reading first line of sam file.
Error: ('SAM line does not contain at least 11 tab-delimited fields.', 'line 1 of file input_bam_file')
[Exception type: ValueError, raised in _HTSeq.pyx:1220]

I am not certain if the problem is with STAR generated bam file or with htseq-count ?
Any help is appreciated

P.S: this is the STAR command given to create the bam file

STAR --genomeDir genome_Dir/genome_mm10 --runThreadN 8 --readFilesIn sample_file --outFilterIntronMotifs RemoveNoncanonical --outSAMmode Full --outStd SAM | samtools view -bS - > Sample1.bam

[chadn737]

htseq-count needs a SAM file, not a BAM file. Try it again with a SAM file. This can be generated with:

samtools view -h -o output.sam input.bam

[priya]

htseq-count needs a SAM file, not a BAM file. Try it again with a SAM file. This can be generated with:

samtools view -h -o output.sam input.bam

Thanks it worked..

[sdriscoll]

Also to save time you can pipe that samtools conversion into htseq-count

samtools view <bam file> | htseq-count [options] - <gtf> > countsfile

I forget if you need to write 'stdin' instead of the '-' for piped input. One of the two will work.