I trying to detect widespread splicing defects and amongst other things I ran tophat to find new splice junctions on my libraries (PE, 100 bases):
Afterwards, the junction.bed files were merged and used as input for DESeq2 (condition vs control, 2 replicates). Then I tried to look at some of the significant events in IGV to get some nice sushi plots... and here lies my problem. This event, and there are more, has a clear pattern, being expressed in one condition and not in the other:
However, when I look at that junction in igv the two libraries that have no expression have a high count:
Is this something to do with the way IGV counts reads in that junction, am I missing something, or is it all wrong?
Code:
tophat -p 12 \ -r 200 --library-type fr-firststrand \ --transcriptome-index=$bowtie_tindex \ -o $outputdir \ $bowtie_gindex \ $fastq_R1 $fastq_R2
chr17_24721664_24721852_chr17_24721714_24721802 0 0 264 591
Is this something to do with the way IGV counts reads in that junction, am I missing something, or is it all wrong?