Empty cluster output

[Dongmeng-wang]

Hi,

I'm using the up-to-date script leafcutter_cluster_regtools.py to cluster junctions, but I get zero clusters in the output. The python version is 3.11 and the junctions were extracted with regtools. samtools index ${sample}.waspfilter.outblacklist.bam regtools junctions extract -a 8 -m 25 -M 1000000 -s XS ${sample}.waspfilter.outblacklist.bam -o ${outdir}/${sample}.waspfilter.outblacklist.bam.junc

I got 0 clusters from two test files, the script and these two files look like this:

I have gone through a few threads saying that the strand of ? might be the issue. However, our data is unstranded. Can I delete if strand == "?": continue from the script? Actually, I didn't quite get why the junctions with ambiguous strands should be ignored. I would really appreciate it if you could give me a clue about this issue. Thanks!

All the best, Meng

[YUTINXIN]

I have encountered the same problem as you before, because you used the wrong parameter. You can try replacing - s XS with - s RF or - s FR.