I have some question regarding the ‘paired’ argument and how to use it in MeDIPS (version 1.23.2)
- If ‘paired’ = true, are the arguments ‘extend’ and ‘shift’ then ignored automatically?
- If ‘paired’ = true, how does normalization with rpkm then works? Is it only based on the reads (as the name implicates) or is it the entire paired-end fragment? I just read about FPKM, where the fragment are considered instead of the reads.
- If ‘paired’ = true, is both the first and second mate then imported? Is it the entire fragment that is included in the analysis or just the reads? An example could be when wiggle files are created: in my data I have reads of 49 bp. I set the ‘window_size’ = 50. Is it then the numbers of reads (49 bp) or also fragments (around 200 bp = spanning several windows) that are counted in each window?
One last question, what is the default setting for 'isSecondaryAlignment'? Is there any recommendations on whether to use only primary alignments or not?