[BioC] Limma

[Seraya Maouche]

Dear  Jim, dear Wei,
Thanks for your help, it is not a two color analysis, it is Illumina.

Best wishes
seraya

-----Original Message-----
From: James W. MacDonald [mailto:jmacdon at med.umich.edu] 
Sent: Mittwoch, 6. April 2011 15:55
To: Wei Shi
Cc: Seraya Maouche; Bioconductor at r-project.org
Subject: Re: [BioC] Limma

Hi Wei,

I think you misunderstood the OP. Seraya _didn't_ remove genes that were
only present in one condition. The problem is that the reviewer didn't like
ratios with a zero in the denominator, which is a fair complaint.

I don't do two color analyses, so don't know what the consensus is for
handling logratios where the denominator is really close to zero. Since you
guys do this stuff all the time, perhaps you have some pointers?

Best,

Jim



On 4/5/2011 6:43 PM, Wei Shi wrote:
> Hi Seraya:
>
> 	Genes which are present in one condition but not in the other should
NOT be removed from your analysis. Only those gene which are absent in both
conditions should be filtered out to improve the power to detect
differentially expressed genes. It is very likely that a lot of genes of
biological interest were not included in your analysis results due to the
removal of genes which are present in one condition but not in the other.
Have a look at the case study for processing Illumina BeadChip data in limma
user guide about the probe filtering.
>
> 	Hope this helps.
>
> Cheers,
> Wei
>
> On Apr 6, 2011, at 2:05 AM, Seraya Maouche wrote:
>
>> Dear Prof Gordon, dear Bioconductor members,
>>
>> I have performed gene expression analysis using Limma (Illumina human
>> ref8) comparing two types of cells (referred below as cond1 and cond2).
>> Based on detection call, I filtered out transcripts which are absent 
>> in both types of cells. Transcripts which were expressed only in one 
>> cell type were included in the analysis.
>>
>> I have received the comment below from a reviewer who seems not agree 
>> to calculate fold change for genes expressed only in one condition. 
>> Would it be possible to have your opinion about this.
>>
>> Thank you in advance for your time,
>> S Maouche
>>
>> "There is a little conceptual difficulty related to the cond1/cond2 
>> comparisons for genes that are considered not detected. If a gene 
>> product is absent (0) in one cell then no fold change can be computed 
>> (table 2). I don’t know how to circumvent this difficult except by 
>> saying that the “noise” is considered to reflect low expression. The 
>> terms “not detected” and “not expressed” are often used 
>> interchangeably while this is not the same. Detection is based on the 
>> definition adopted and in many places of the manuscript it should be 
>> used in place of expression."
>>
>>
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--
James W. MacDonald, M.S.
Biostatistician
Douglas Lab
University of Michigan
Department of Human Genetics
5912 Buhl
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