How to make the script work with stringtie command first and then merge the outputs?

Question

[Biologist]

Hi,

With hisat2 command I have bam files as output and sorted them to use as input for stringtie.

I have a bash script like below. First it will take sorted.bam files as input and give gtf as output. Then path for each sample gtf will be given into merge list.txt. and then use stringtie merge on them.

I totally have 40 sorted.bam files.

for sample in /path/*.sorted.bam
do
  dir="/pathto/hisat2_output"
  dir2="/pathto/folder"
  base=`basename $sample '.sorted.bam'`
  "stringtie -p 8 -G gencode.v27.primary_assembly.annotation_nochr.gtf \
    -o ${dir2}/stringtie_output/${base}/${base}_GRCh38.gtf \
    -l ${dir2}/stringtie_output/${base}/${base} ${dir}/${base}.sorted.bam; 
  ls ${dir2}/stringtie_output/*/*_GRCh38.gtf > mergelist.txt; 
  stringtie --merge -p 8 -G gencode.v27.primary_assembly.annotation_nochr.gtf \
    -o ${dir2}/stringtie_output/stringtie_merged.gtf mergelist.txt"
done

I separated the commands with ; After running the script on all sorted.bam files and after completing the job I see that mergelist.txt has paths only for 33 sample gtf's. Which means the path for other 7 sample gtfs is missing in merge list.txt.

How to make the script work with one command first and then to next command?

[cschu181]

Maybe run stringtie manually on one or two of the missing samples and check if that generates anything or whether it fails for those samples.

On a side note, I would move the ls ${dir2}/stringtie_output/*/*_GRCh38.gtf > mergelist.txt; and stringtie --merge -p 8 -G gencode.v27.primary_assembly.annotation_nochr.gtf -o ${dir2}/stringtie_output/stringtie_merged.gtf mergelist.txt out of the loop. You only want to do that once after all samples have been processed. You can also move the dir="/pathto/hisat2_output" and dir2="/pathto/folder" to be executed before the loop, as they don't seem to be sample-specific.

[Biologist]

Is this what you want to me to change in the script?

  dir="/pathto/hisat2_output"
  dir2="/pathto/folder"
for sample in /path/*.sorted.bam
do
  base=`basename $sample '.sorted.bam'`
  "stringtie -p 8 -G gencode.v27.primary_assembly.annotation_nochr.gtf \
    -o ${dir2}/stringtie_output/${base}/${base}_GRCh38.gtf \
    -l ${dir2}/stringtie_output/${base}/${base} ${dir}/${base}.sorted.bam"
done

  ls ${dir2}/stringtie_output/*/*_GRCh38.gtf > mergelist.txt; 
  stringtie --merge -p 8 -G gencode.v27.primary_assembly.annotation_nochr.gtf \
    -o ${dir2}/stringtie_output/stringtie_merged.gtf mergelist.txt

[cschu181]

Yes, but make sure to check what stringtie does with the bam files for which you don't get output. The script changes should only affect run-time.

[Biologist]

Yes, when ran the stringtie with bam files which I don't get in mergelist.txt works when I gave manually.

[Biologist]

In the above way it gives an error. I submitted the job.

ls: cannot access /path//_GRCh38.gtf: No such file or directory Error: no transcripts were found in input file mergelist.txt

[cschu181]

Remove the "" around "stringtie -p 8 -G gencode.v27.primary_assembly.annotation_nochr.gtf \ -o ${dir2}/stringtie_output/${base}/${base}_GRCh38.gtf \ -l ${dir2}/stringtie_output/${base}/${base} ${dir}/${base}.sorted.bam" ?