What Tools Are Available For Analyzing Bisulfite Data?

Question

[Christopher Bottoms]

A colleague produced an alignment (SAM format) of bisulfite-treated sequences using Novoalign. About 85% of the sequences seem correctly aligned (which may need to be resolved first). Is there a tool or set of tools available for finding and quantifying the C->T conversions? The read length for each aligned sequence is roughly 40bp.

I'm most familiar with Perl, but I'm open to tools in any language that would be helpful.

[Christopher Bottoms]

@Pierre Lindenbaum. Where did your answer go?

Answer 1

[brentp]

I had that same problem with existing pipelines for BS-treated reads--it's hard to get per-base methylation scores after the alignment.

So, (shamelss plug) I wrote MethylCoder, which takes FASTQ or Fasta reads files runs the alignment, and outputs (in addition to a SAM file) a text file like:

#seqid  mt  bp  c   t
chr1   3   1354    0   1
chr1   1   1358    6   1
chr1   2   1393    0   1
chr1   6   1394    4   9

indicating the methylation status at every base-- the methylation is c / (c + t).

where mt is the methylation type or context (1/4 are CG +/- strand, 2/5 are CHG +/- strand and 3/6 are CHH +/- ).

The pipeline is documented at the link above and also includes scripts for visualization and analysis for doing things like finding regions that are differentially methylated between 2 runs of BS-Seq. Also the author is very responsive to questions and feature requests ;)

Answer 2

[Eric Fournier]

I understand that this question was answered ages ago, but since it is the most relevant question tagged "Bisulfite" and thus a natural first step for those new to the technology and seeking general guidance on how to interpret it (as I did a month ago), I think the following update is relevant:

Felix Krueger wrote the following review in early 2012 detailing the steps for analysis of bisulfite-treated libraries, common sources of errors and a list of tools. Among these tools are Bismark(ref) (written by Krueger), MethylCoder (ref) (written by brentp), BRAT (ref), BS-Seeker (ref) and BSMAP (ref).

I'd also recommend going to the SeqAnswers software list and searching for "bisulfite"/"bisulphite".

Answer 3

[Daniel Standage]

I've been very pleased with the GNUmap software and the response of its developers. They have a flag for doing bisulfite analysis. However, I'm not sure if you can do the bisulfite analysis on an existing alignment, you may have to re-map the reads if you use this tool. But definitely for the future I would recommend this tool.

Answer 4

[Fred Fleche]

[?]

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http://quma.cdb.riken.jp/

Answer 5

[Alastair Kerr]

There were new tools advertised in the ISMB this year. I have not tried them myself but search the proceedings Especially the poster sessions.

[Christopher Bottoms]

e.g. Bismark(http://seqanswers.com/forums/showthread.php?t=5502)?

[Christopher Bottoms]

GNUMAP (mentioned by Daniel Standage) was also presented in a poster.

Answer 6

[Neilfws]

You may want to look at methVisual, a Bioconductor package and BSMAP, a tool to map bisulfite reads to genome sequence. Disclaimer: I have not used either, so can't be sure of their suitability for your problem.

Answer 7

[brentp]

As another answer, GSNAP (part of GMAP) maps BS-Seq reads. In my experience, it is able to map more reads, at the expense of running time.

There is a comparison of what I consider the full-featured BS-Seq mapping programs here: https://github.com/brentp/methylcode/tree/master/bench

Answer 8

[xieshaojun0621]

Hope ViewBS can help you. https://github.com/readbio/ViewBS