Hello everyone, I'm trying to use stringtie to quantify some previously mapped RNA-seq reads. And I have some basic questions. I appreciate if someone explains to me in simple words:
How does certain quantifier's can only quantify the output of spliced aligners? Does it make any difference? After all, these are all mapped reads, what is specific about spliced vs non-spliced aligner outputs?
In case of stringtie and i'm sure many other qunitifiers, there is an option for specifying the strand, e.g.: --rf Assumes a stranded library fr-firststrand. --fr Assumes a stranded library fr-secondstrand.
first of all, what is a library? Is it the very first set of raw reads? Eventually, each read will map to one strand, so we will know the strand, is it not? What am I missing?
And my last question is about the XS tags, why does it matter only when we are talking about the junctions? Thanks