bam file sorted by "-n" was not accepted by qualimap

Question

[Yingzi Zhang]

Hey guys, I was using qualimap to estimate the features of a .bam file. I input the .bam file and it reported like this:

Failed to run bamqc java.lang.RuntimeException: The alignment file is unsorted. Please sort the BAM file by coordinate.

My raw data are hiseq 2000 100bp x 2 short reads, I thought I could only sort them by using "-n" parameter. So I sorted them by samtools sort -l 9 -o output -n -@ 40 input Then I used qualimap like this qualimap --java-mem-size=700G bamqc -bam .mem.srt.bam -gff <dirpath/.gff> -oc .count.matrix -outdir </bamqc> -outformat PDF:HTML Is my code wrong? how should I fix it please? Thank you.

[WouterDeCoster]

Hey guys

And what about non-guys? Please use neutral words :)

[Yingzi Zhang]

okay, thanks for reminding :P

Answer 1

[GenoMax]

Then try without the -n parameter. Name sorted files are only needed for some specific purposes (if you were converting them to fastq for example). Normally BAM files are co-ordinate sorted and that is how qualimap wants them for producing stats.

On a different note: you are allocating 700G for a qualimap run? Are your files TB+ is size?

[Yingzi Zhang]

It worked fine without the -n. Thank you very much. I allocated 700G for the both situations because I don't know how much space java need. So just kind of in case. Would it be stupid? I later found I should do multi-bamqc. The bam files are 324G in total. And still I allocated 700G. Any advice please?