Hi Jonathan,

CpG density normalization of MeDIP-seq data is still possible using MEDIPS
v >= 1.10.0: by setting the parameter MeDIP of the MEDIPS.meth() function
to MeDIP=TRUE, the resulting table will contain an additional column for
CpG density normalized rms values per sample.

In principle, the concept of CpG density normalization remains the same and
the dependency between CpG density and read coverage can still be examined
by plotting the calibration plot. However, there are several conceptional
modifications in the new version that will result in differences when
comparing results of the original and of the updated version. For example,
read coverage and CpG density normalization is performed directly on the
targeted window size (e.g. genome wide 300bp windows) and not at smaller
(50bp) bins any more. Consequently, calculation of ams values became
obsolete. In section 7.7 of the manual, I briefly describe concerns I have
today with deducing percentage methylation from CpG density normalized rms/
ams values.

In summary, i) I recommend to use rms values for comparisons to bisulfite
data, ii) I consider it to be problematic to deduce percentage methylation
from the rms values without additional knowledge (see section 7.7 of the
manual), and iii) I would like to emphasize that the main focus of the new
MEDIPS package is clearly on the identification of genome wide differential
methylation (or differential coverage in general) between groups of samples.

All the best,
Lukas



On Thu, Aug 15, 2013 at 6:02 PM, Jonathan Ellis
<Jonathan.Ellis@qimr.edu.au>wrote:

> Dear Lukas and Bioconductors,
>
> I have been asked to look at a MeDIP-Seq dataset that has been processed
> with a previous version of the MEDIPS package (version 1.2.0) and I've
> noticed there have been some changes with regards to the current version
> I have installed (version 1.10.0).  In particular, it appears as though
> the whole concept of AMS (absolute methylation score) has been dropped
> from the most recent version.
>
> The previous analysis of the data I'm currently looking at used AMS
> values, and I hoping someone can tell me why it's no longer used (e.g.,
> is it no longer considered a useful metric, or is there something else
> that better describes the methylation?).
>
> The 1.2.0 version documentation contains the following (on page 27):
>
> "We have shown that such summarized methylation values for ROIs,
> especially the ams values, are most suitable for comparing MeDIP data to
> e.g., bisulhpite sequencing or whole genome shotgun bisulphite
> sequencing data"
>
> If this is correct, what is the best metric to use for such comparisons
> now ams values are no longer being produced?
>
> Thanks,
> Jonathan
>
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