I have stranded RNA-seq data which doesn't look stranded when I visualize bedGraphs in Genome Browser:

![enter image description here][1]

I tried different aligners (star, bowtie, bwa), different ways to make bedGraph files, nothing works. But when I run salmon on these samples, it detects ISR (paired-end stranded library). I'm very confused why it doesn't look stranded in the browser but I know is stranded. Any help will be really appreciated! I've been trying to solve it on my own for a while...


  [1]: https://s8.postimg.cc/4yxzbs4bp/strand_error.png
Hi,

  The reason for this is that whether or not to consider a fragment for quantification in salmon is considered *after* the mapping is written.  That is, the reads are mapped to the transcriptome, without regard for the inferred or provided library type, and then these mappings are written to file if requested by the user.  However, when salmon considers the probability of these mappings arising from different transcripts, it will discard those that are not compatible with the library type.  So, my guess is that you're seeing in this output multi-mapping reads where the sequence matches, and hence there is a possible allocation of this read to the transcript, but for which these mappings are later discarded during quantification.  You could, for example, filter the mapping file based on the tags of the alignment records, to only keep those reads that align according to the ISR library type.