[BioC] Interspecies differential expression of orthologs with Edger

Gordon K Smyth smyth at wehi.EDU.AU
Sun Sep 7 03:11:00 CEST 2014

Dear Assaf,

You are getting the sort of results that I would expect you to get when 
you try to compare two RNA sources that are very different.

The diagonal lines in the MA plot are simply a result of having low counts 
(0,1,2 etc) in one species and high counts in the other for the same 

When you compare different species, I'd intuitively expect almost every 
gene to be differentially expressed to some degree.  So I'm not surprised 
that a large proportion of genes are assesssed as DE.

That's about as much help as I can give you.  I can't give advice that 
would allow you to get the same sort of results as you might be used to, 
because comparing different species isn't a normal thing to do.

Best wishes

> Date: Fri, 5 Sep 2014 23:22:28 +0300
> From: assaf www <assafwww at gmail.com>
> To: Gordon K Smyth <smyth at wehi.edu.au>
> Cc: Bioconductor mailing list <bioconductor at r-project.org>
> Subject: Re: [BioC] Interspecies differential expression of orthologs
> 	with	Edger
> Thanks Gordon,
> To summarize the results I got on the cross-species data, after embedding
> the length-effect to the GLM offset matrix, as in the code you sent, please
> see the attached MA plot:
> 1) for >5 and <-5 log fold change, genes' logFC is positively correlated
> with mean log CPM, something I haven?t seen before in Edger standard runs.
> 2) most genes with fold change around > 1.3, or < -1.3, are significant,
> which looks to me too ?liberal?. Please note that each group contains 6
> true biological replicates (variance within each group is large) .
> The first problem worries me most, any idea is very welcomed.
> Many thanks,
> Assaf
> On Wed, Sep 3, 2014 at 2:08 AM, Gordon K Smyth <smyth at wehi.edu.au> wrote:
>> On Tue, 2 Sep 2014, assaf www wrote:
>>  Does Edger DE analysis is built on the assumption that most genes are not
>>> differentially expressed, and that only a small portion of them do (say
>>> <20%)  ?
>> Only the calcNormFactors() step of edgeR makes any assumption of this
>> sort. calcNormFactors assumes that either that most genes are not DE or
>> that the DE is reasonably symmetric.
>>  I mean, in cross-species studies, or when comparing different tissues of
>>> the same organism, if this assumption doesn't hold, should it be a serious
>>> concern ?
>> In a cross-species comparison there will be many DE genes, but some will
>> be up and some will be down.  The DE will not be all in one direction, I
>> would guess that normalization will not be a serious concern.
>> Of all the concerns with cross-species comparisons, this seems to me to be
>> far from the most serious.
>> Best wishes
>> Gordon
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