[BioC] Limma contrasts question

[Daniel Brewer]

James W. MacDonald wrote:
> Daniel Brewer wrote:
>>
>> Thanks for that information.  Yes I did mean PC3M as a contol ... doh.
>>
>> Unfortunately I am still having problems.
>>> contrast.matrix <- makeContrasts("KvC" = Knockdown - (Scramble +
>> PC3M)/2, levels = design)
>> Error in eval(expr, envir, enclos) : object "PC3M" not found
>>
>> I believe this is because PC3M is not in the design matrix as I am using
>> it as the reference.  As a structure for this I have been using the
>> direct 2-color design example of the "Limma User guide" (Section 7.4).
>> There is a discrepancy between the straight comparisons that you have
>> given me and the example in the guide.
>>
>> Your suggestion:
>> contrast.matrix <- makeContrasts("KvsC" = Knockdown - PC3M, levels =
>> design)
>> Their suggestion:
>> contrast.matrix <- makeContrasts("KvsC" = Knockdown, levels = design)
>>
>> I think this will have something to do with PC3M being described as the
>> reference (the intercept in the linear model???) in their approach but
>> maybe you use a different reference.  Not sure.
> 
> Oh yeah. Sorry - I didn't look closely at the design matrix. I don't do
> two color stuff so this doesn't come second nature to me.
> 
> Anyway, I think your original contrasts matrix should be OK.
> Algebraically (2*Knockdown - Scramble)/2 should equal Knockdown -
> (Scramble + PC3M)/2. Sorry for the noise.
> 
> 
> Best,
> 
> Jim
> 
> 
> 
>>
>> Many thanks for your time.  I really would like to get an understanding
>> of this.
>>
>> Thanks
>>
>> Dan
>>
> 

Hi Jim,

Could you go into the maths of the contrast formulas a bit?  I would
like to get a really solid understanding of what it is doing for future
analyses.

Many thanks

Dan

-- 
**************************************************************

Daniel Brewer

Institute of Cancer Research
Molecular Carcinogenesis
MUCRC
15 Cotswold Road
Sutton, Surrey SM2 5NG
United Kingdom

Tel: +44 (0) 20 8722 4109
Fax: +44 (0) 20 8722 4141

Email: daniel.brewer at icr.ac.uk

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