Missing major exon skipping event

[Chris Seward]

I've been experimenting with Majiq Build v2.2-e25c4ac to determine if I understand how to use it.  

I have a test dataset from two mouse strains, one of which has a knockout of two exons in a gene.   We have 9 RNASeq samples for each genotype, with a sequencing depth of 40-50m, paired end, 150bp reads per sample, mapped to the mm10 ensemble genome and ensemble v102 annotation with STAR.

MAJIQ does find a few (~10-20) very minor (hard to spot visually) LSV between the two strains, but they are only located in non-standard chromosomes like chrX_random, chrUn_JH584304 etc.  Does this indicate something is off?  What is a normal range of numbers of LSV you would expect to find in an actual animal experiment?  100s? 1000s?

I am wondering if MAJIQ is ignoring the actual standard chromosomes for some reason?  In particular, it never identifies the actual exon knockout we engineered as a variant.  

I can see the variant very clearly myself in the bam files, but it just isn't being spotted computationally.  Do you think I am doing something wrong or is this something the program normally wouldn't detect for some reason?

[Chris Seward]

After more troubleshooting I've determined it was an issue with the genome annotation I was using.  I think it was a "chr1" vs just "1" problem (ucsc vs ensemble), but now I've got it working and am finding the top hits I expected to find!  It was only working before on the non-standard chromosomes because the nomenclature for those chromosomes is the same between the annotations.  Posting this here in case other people have similar issues!

[Paul Jewell]

Hello,

I'm glad you were able to work out part of the issue. I wanted to mention that there was also a separate longstanding issue people were having with specifying specific strandness in the majiq config file, finding many fewer LSVs than intended. We had a temporary fix for this released in a development branch, but I have just merged it into the master branch today. You can try to reinstall if you were using the strandness specification other than "none" in your config file.

Thanks.