I am for the first time doing a TE analysis using Ribo-seq and RNA-seq data, however I have a few question regarding the analysis.
I have used STAR to align the reads from both datasets to the Human genome. For the RNA-Seq datatsets (average read length 100 nt) I get a very good amount of uniquely mapped reads. For the Ribo-seq datasets I get only 13-15% of uniquely mapped reads, which I guess is understandable if we consider that the average read length is very short (30 nt).
I am now planning to get the gene counts (as TPM) for both datasets and use these to calculate the translation efficiency of each gene. Should I use all mapped reads independently on the fact that most of them show multi-mapping or should I just use the uniquely mapped reads? In the second case, won't this be underepresentative of the real counts?
Thanks a lot in advance for your help.