[BioC] BioC normalisations for small array 2 colour data?

[Gordon Smyth]

Hi Naomi,

I agree, spike-in can't be relied on for normalization. I had in mind 
pooled titration controls as in Yang et al (2002), which can be 
printed on the arrays but which don't need to be spiked into the RNA samples.

Best wishes
Gordon

Yang, Y. H., Dudoit, S., Luu, P., Lin, D. M., Peng, V., Ngai, J., and 
Speed, T. P. (2002). Normalization for cDNA microarray data: a robust 
composite method addressing single and multiple slide systematic 
variation.  Nucleic Acids Research 30(4):e15.

At 11:32 PM 8/09/2006, Naomi Altman wrote:
>Spiking in normalization control probes can also be problematic, as 
>the RNA samples have to be equalized before spiking.
>This can, I think, be done when the tissues from which the RNA is 
>extracted are very similar, and the extraction protocol is very
>uniform.  Otherwise, spiking variation just gets added to all the 
>other sources of variation.
>
>--Naomi
>
>At 08:07 AM 9/8/2006, Gordon Smyth wrote:
>>Dear Dan,
>>
>>At 08:00 PM 8/09/2006, bioconductor-request at stat.math.ethz.ch wrote:
>> >Date: Thu, 7 Sep 2006 13:44:35 +0100
>> >From: "Dan Swan" <bioinformatics.lists at gmail.com>
>> >Subject: [BioC] BioC normalisations for small array 2 colour data?
>> >To: bioconductor at stat.math.ethz.ch
>> >
>> >Hi,
>> >
>> >I have some data from a small specialised microarray - 200 genes, 1
>> >spiked control, 1 negative control.  This is 2 colour data, with dye
>> >swaps.  I was wondering what an appropriate normalisation for this
>> >scenario is within Bioconductor given that Lowess is unreliable for
>> ><1000 genes?
>>
>>Are you quoting someone when you say that lowess is unreliable for
>><1000 genes? Who? Print-tip loess normalization is routinely applied
>>to arrays with around 200 genes in each print-tip group so 200 genes
>>is, far from being unusual, pretty much typical for loess normalization.
>>
>>The real problem with a specialised microarray is the fact that the
>>genes are not randomly chosen, indeed they are typically chosen
>>because of their likelihood to be differentially expressed. Hence you
>>may be in a situation where the majority of genes may be
>>differentially expressed. If that is your case then, in my opinion,
>>normalization control probes need to be designed into the array in
>>the first place to produce reliable results.
>>
>>Best wishes
>>Gordon
>>
>> >Any pointers would be gratefully recieved.
>> >
>> >thanks,
>> >
>> >Dan
>> >
>> >--
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>
>Naomi S. Altman                                814-865-3791 (voice)
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