Hi everyone! I'm performing some ChIP seq experiments and I want to read depth coverage over the region -500 to 3000pb of the TSS of all Ensemble protein coding transcript.
I have different approach, but as I am totally naive, I appreciate very much your advises.
a)First of all I've normalized the bam files using bamCoverage (deepTools), the output is a bedgraph, then I will do the anntotaion peak over TSS using ChIPseeker (R)
b)Also, I can do the peakannot over the bed file (after bam file conversion) and I will normalize the peakannot when I will plot it.
Other questions is, Is necessary make the peakcalling to calculate the read depth over TSS?
I'm looking forward to hear from you
Than you!!!
Hi again, As ChIPseeker gives me some error, you know any other way in which I can get the read depth coverage over TSS onces I have the normalized file ? Thanks!!
There are many many tools to do that. Since you are already familiar with deeptools, you could try their computeMatrix tool.
Thank you, Do you know what file can I use as TSS.bed in computeMatrix? I downloaded one from USC but the program gives to me an error message
Warning: chrY:9748406-9748406 is an invalid BED interval! Ignoring it.
Thanks!
Could it be that your ChIP-seq data is from a female and that there is no Y ? This is a warning, not an error, and it could be ok to proceed if the non-Y coordinates are considered correct by the script.
More generally, check that the chromosome names in the bed file are the same as in the bedgraph file.
Carlo