Methylation Extractor report missing Chr annotation

Question

[Jonathan Crowther]

Hi there,

So I have recieved some Bismark output files:

Bismark (SAM/BAM)

Mbias files & Plots

Methylation Extractor reports

The documentation is very good and I have a good understanding of all the files however in my data from the Methylation extractor there are several sites that are not marked with the chromosome header.

The report files according to methylationExractor should have the following headers:

(1) seq-ID (2) methylation state (3) chromosome (4) start position (= end position) (5) methylation call

In my files I have as follows:

HWI-ST571:431:C43FMACXX:8:2316:9928:82689_1:N:0:GTCCGC  -       chr9    23913539        z
HWI-ST571:431:C43FMACXX:8:2316:9928:82689_1:N:0:GTCCGC  +       chr9    23913531        Z
HWI-ST571:431:C43FMACXX:8:2316:9928:82689_1:N:0:GTCCGC  +       chr9    23913529        Z
HWI-ST571:431:C43FMACXX:8:2316:9928:82689_1:N:0:GTCCGC  -       chr9    23913485        z
HWI-ST571:431:C43FMACXX:8:2316:9928:85484_1:N:0:GTCCGC  +       chr2    28594224        Z
HWI-ST571:431:C43FMACXX:8:2316:9928:85484_1:N:0:GTCCGC  -       chr2    28594341        z
HWI-ST571:431:C43FMACXX:8:2316:9928:85757_1:N:0:GTCCGC  -       30126    6594       z # <- this line
HWI-ST571:431:C43FMACXX:8:2316:9928:85757_1:N:0:GTCCGC  -       chr2    223724995       z
HWI-ST571:431:C43FMACXX:8:2316:9929:12206_1:N:0:GTCCGC  -       chr12   64474195        z
HWI-ST571:431:C43FMACXX:8:2316:9929:59562_1:N:0:GTCCGC  +       chr20   55837341        Z
HWI-ST571:431:C43FMACXX:8:2316:9929:59562_1:N:0:GTCCGC  -       chr20   55837591        z
HWI-ST571:431:C43FMACXX:8:2316:9929:59562_1:N:0:GTCCGC  +       chr20   55837557        Z
HWI-ST571:431:C43FMACXX:8:2316:9929:59562_1:N:0:GTCCGC  +       chr20   55837519        Z
HWI-ST571:431:C43FMACXX:8:2316:9929:89987_1:N:0:GTCCGC  +       chr5    92603043        Z
HWI-ST571:431:C43FMACXX:8:2316:9929:89987_1:N:0:GTCCGC  +       26965    45121        Z #<- this line
HWI-ST571:431:C43FMACXX:8:2316:9929:89987_1:N:0:GTCCGC  +       26965    301        Z #<- this line
HWI-ST571:431:C43FMACXX:8:2316:9929:89987_1:N:0:GTCCGC  +       chr5    92603089        Z
HWI-ST571:431:C43FMACXX:8:2316:9929:89987_1:N:0:GTCCGC  +       chr5    92603122        Z

I am not sure about this as they seem to be a strange format. Initially I though maybe there was a formatting issue and it skipped the "chr" but none of the numbers correspond to a CG site.

When I run bismark2bedgraph the job completes and these files are also listed but without the chromosomal location I cannot locate the methylation site.

Have any of you observed this before or know what has happened?

Cheers

[Devon Ryan]

That looks like a bug in the program. Felix, the author of bismark, isn't particularly active here but is on seqanswers. You might just post this as a bug report on the bismark thread over there. In the interim, if you convert your alignments to a BAM file and sort/index them, you can just use PileOMeth to directly make the bedGraph files.

[Devon Ryan]

For those coming across this later, there's a follow-up on SEQanswers.