Hi!
I have a set of .BAM files (with reads) on different histone marks but I do not have replicates.
The files that I have are H3K27ac (untreated), H3K27ac (treated with sample1) and H3K27ac (treated with sample2).
I have already called peaks on these samples using MACS2.
I want to identify differential peaks across these three.
Questions:
a) How can I identify these peaks?
b) Is their any package available to do so, if yes, have you used it?
Kindly help.
Thank you
4 answers
I highly recommend you to try diffreps http://code.google.com/p/diffreps/
yes, it works without replicates, but you need to use G-test
With no replicates, you might be best off just overlapping the peaks found and separating it into classes of overlaps (think venn diagram). You could also give DBChIP a try ( bioconductor link ) since it has an alternate way that tries to estimate variation w/out replicates.
Please checkout the RSEG tools at http://smithlab.usc.edu/histone/rseg/ I developed this tool using Hidden Markov model. If you use the rseg-diff program with mode 3, it will identify regions with differential read counts.
Please let me know if you have any questions.
These guidelines might be helpful:
Bailey T, Krajewski P, Ladunga I, Lefebvre C, Li Q, Liu T, Madrigal P, Taslim C, Zhang J (2013) Practical Guidelines for the Comprehensive Analysis of ChIP-seq Data. PLoS Comput Biol 9(11): e1003326. doi:10.1371/journal.pcbi.1003326
http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003326
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If you're hoping to get reliable p-values, you won't get any without replicates. You might just make a consensus peak set and then calculate pairwise fold-changes of normalized coverage between the conditions in them.