Question: Calculate the total genie length targeted in my chIP
0
gravatar for Zee_S
3 months ago by
Zee_S20
Zee_S20 wrote:

Hello Everyone,

I have a list of annotated meta genes in a bed file. I also have my chIP.bam and INPUT.bam files. How do I calculate the total length of basepairs lying within these genes that are enriched for my chIP?

Thank you for your support!

ADD COMMENTlink modified 3 months ago by zx87544.7k • written 3 months ago by Zee_S20
2
gravatar for Devon Ryan
3 months ago by
Devon Ryan81k
Freiburg, Germany
Devon Ryan81k wrote:

You would need to call peaks and then see how much they overlap (e.g., with bedtools intersect or bedtools coverage).

ADD COMMENTlink modified 3 months ago • written 3 months ago by Devon Ryan81k

Hello Devon,

Thanks a lot for your reply. I m looking at a broadly distributed mark so I don't call peaks. I usually compute a log2ratio of fold enrichment over input and use this to define my regions. Is it correct to say that using bedtools coverage, if I sum up the total number of bp per gene that have a log2ratio >=1.0 for example, is my enriched genic length ?

Many thanks

ADD REPLYlink written 3 months ago by Zee_S20
1

Possibly, though I would encourage you to use a domain caller for broad marks rather than using a simple log2 ratio.

ADD REPLYlink written 3 months ago by Devon Ryan81k

Hello Devon, Thank you for your reply. Would you have any suggestion of a suitable way to get a log2 ratio when the input has zero reads assigned? Assuming I have comparable coverage in both IP and Input, can I assign a pseudo count of +1 to these "zero reads" bins? my worry is that this would change the ratios significantly and affect my downstream calculations of gene length.

even if I use a domain caller, as you recommend, this problem still persists (I think)

Thanks again.

ADD REPLYlink written 3 months ago by Zee_S20

As you surmised, you need to add a pseudo-count. I think we default to 1 for this in deepTools, I haven't seen any better suggestion for that value.

ADD REPLYlink written 3 months ago by Devon Ryan81k
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