Merging .bam files in STAR

Question

[sarah.laverty20]

Hi there,

I have successfully created an annotated genome index to map my paired-end rna-seq reads and have run:

STAR --runMode alignReads --genomeDir /home/rnaseq/ref/ --outSAMtype BAM SortedByCoordinate --readFilesIn R21_1.fq.gz R21_2.fq.qz --runThreadN 10 --outFileNamePrefix /home/rnaseq/fastq/mapped/R21 --readFilesCommand gunzip

I believe in order to get one .bam file from my paired-end reads I insert both file names with a space between after --readFilesIn. When I run this I receive this error message:

!!!!! WARNING:  Could not ls R22_1.fq.qz
EXITING: because of fatal INPUT file error: could not open read file: R22_2.fq.qz
SOLUTION: check that this file exists and has read permision.

I have played around with the files I am using and their order and it seems that no matter what file it is, STAR is unable to read the second file listed. If I run them individually they map successfully so I don't think there is anything wrong with the file itself.

Any help would be really appreciated as this is my first time processing rna-seq data!

[brunobsouzaa]

Try using --readFilesCommand zcat instead of gunzip.

In this case, you won't need to unzip your files!

[sarah.laverty20]

That has worked, thanks so much!

[Pierre Lindenbaum]

what is the output of ls -lah R21_1.fq.gz R21_2.fq.qz

[sarah.laverty20]

I believe it is because my --readFilesCommand gunzip -c command was only unzipping the first file and not the second and therefore the second .fq.gz file was unreadable.

I have now unzipped both files and run the command without --readFilesCommand and it has worked. However, I don't want to have to do this for all of my samples.

Do you know how I can tweak the command to unzip both files?