Hi,
Any answers would be highly appreciated as I've been stuck on this for a while. I analyze epigenetic data and process DMR lists and I'm currently having issues producing a pie chart that shows the distribution of the repeat sequences in my samples. I've already done this for genomic distribution (TSS, 5' UTR etc.) but I can't seem to be able to figure out how I can use RepeatMasker to do the same thing.
Here's exactly what I did for the genomic distribution pie chart:
peakAnno <- annotatePeak("~/Desktop/1-m1Vsm2.bed.gz", tssRegion=c(-3000, 3000), TxDb=txdb, annoDb="org.Mm.eg.db")
plotAnnoPie(peakAnno)
The results looked fine but then I tried the following using the UCSC RepeatMasker package in R (mm39 mouse sample). I've read the documentation and everything and still haven't figure it out. I got the following error:
#Annotate peaks (with repeat sequences) & plot
ah <- AnnotationHub()query(ah, c("RepeatMasker", "Mus musculus"))
rmskmm39 <- ah[["AH99013”]]
peakAnnoRepeat <- annotatePeak("~/Desktop/1-m1Vsm2.bed.gz", tssRegion=c(-3000, 3000), TxDb=txdb, annoDb="rmskmm39")
plotAnnoPie(peakAnnoRepeat)
Warning messages:
1: In library(package, lib.loc = lib.loc, character.only = TRUE, logical.return = TRUE, :
there is no package called ‘rmskmm39’
2: In getGeneAnno(annoDb, peak.gr$geneId, type, columns) :
ID type not matched, gene annotation will not be added..
I thought extracting the mm39 masked repeat track would act as the annotation track instead of "org.Mm.eg.db" as was used in the original working example. If anyone has any pointers or solutions, that'd be great.
Thank you
Wouldn't you need to make a custom TxDb object? I haven't done it myself, but I understood the annoDb to basically map gene annotations to genes given by txdb. Meaning the "peaks" are annotated using txdb info, then the additional gene info (SYMBOL, description etc) comes from annoDb.
I thought that's what I was doing by:
It was my understanding that the rmskmm39 variable was now the repeat annotation track, I can't find a package that includes the repeat sequences like the non-repeat sequences anno track
What is in your txdb object?
Initially, when it worked:
When trying to get the repeated sequences distribution (rmskmm39 variable can be found in original post):
When I tried this, chipseeker ignores the annoDb since it was a "custom" txdb. However it also wouldn't run plotAnnoPie on the resulting chipseeker object, but the regions were annotated with the info from the rmsk table and it would be straightforward to make a pie chart yourself from the columns.
Do you mind telling me how you were able to create a custom txdb object that sufficiently annotates repeats?
I believe I essentially followed your workflow with rmskmm39 as the txdb object.
But, it doesn't work with plotAnnoPie, so you can convert the chipseeker object to a dataframe
Also, you may need to exclude your annoDb argument, since it will be ignored and the "rmskmm39" package isn't valid which may cause the function to fail.
You should then be able to explore the table and manually count up the categories you are interested in.
P.S. I realize now I used the mm10 version of rmsk since that's what I used for my peaks. This probably shouldn't matter though.
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Fixed it, do you happen to have an idea for how to resolve my issue?
Sorry, I'm not equipped to help you with that - I'm not familiar with repeatmasker. I'll clean up these comments and bump the post, that way others will be able to help you.