Problem with methylKit and Bismark

Question

[ahmad mousavi]

Hi

I have used Bismark for aligning RRBS reads and now I should run methylKit on bismarks' output, like CpG and bedGraph and .Cov. How can I use these files instead of bam file.

Methylkit manual said it accepts bismark bam file however, I could not handle it. It gives error when I used bismark sorted bam files, and methRead function does not work too.

Does anybody know pipeline for working with bismark output files? I don't mean bam/sam.

[igor]

Which files did you use the for methRead function?

If it gives you an error, can you post the command?

Answer 1

[jomo018]

Two step solution: Use coverage2cytosine to convert .cov file to an intermediate file. Then a one-liner awk command or some other script to convert the intermediate file to a methRead input file.

The reason for this two-step process is that .cov lacks strand information.

cov file format:             chr  pos  pos  %meth  #meth  #unmeth
intermediate file format:    chr  pos  strand(+/-)  #meth  #unmeth   na  na 
methRead input file format:  id   chr  pos  strand(F/R)   [#meth+#unmeth]  %meth %unmeth

Where # is (integer) count. The intermediate file contains CG sites not covered by any read and you should discard those in your script.

Answer 2

[xuanli1006]

Hi

I have created a file as methylKit input file; it likes the methylKit's test and control samples: chrBase chr base strand coverage freqC freqT 1 chr21.9764539 chr21 9764539 R 12 25.00 75.00 2 chr21.9764513 chr21 9764513 R 12 0.00 100.00 3 chr21.9820622 chr21 9820622 F 13 0.00 100.00 4 chr21.9837545 chr21 9837545 F 11 0.00 100.00 5 chr21.9849022 chr21 9849022 F 124 72.58 27.42

But when I used methRead to read the file, I got errors from this step: Exception: Argument 'file' should be a single value not 7 values. at #17. getVector.Arguments(static, s, length = length, .name = .name) - getVector.Arguments() is in environment 'R.utils'

at #16. getVector(static, s, length = length, .name = .name) - getVector() is in environment 'R.utils'

Error: Argument 'file' should be a single value not 7 values.

Can you help to solve this problem? Thank you very much!

[jomo018]

Suggest you show the script lines preceding and including methRead. Please Format the lines with the proper icon.

[xuanli1006]

I have solved this problem. Thank you for your reply!