[BioC] Differential expression in time series

[James W. MacDonald]

Hi Lev,

Lev Soinov wrote:
> Dear List,
>   On the page 49 of the LIMMA user guide there is an example of how to make a difference of differences in experiments with several factors:
>    
>   "Which genes respond differently in the mutant relative to the wild-type?
>   > cont.dif <- makeContrasts(
>   + Dif6hr =(mu.6hr-mu.0hr)-(wt.6hr-wt.0hr),
>   + Dif24hr=(mu.24hr-mu.6hr)-(wt.24hr-wt.6hr),
>   + levels=design)
>   > fit2 <- contrasts.fit(fit, cont.dif)
>   > fit2 <- eBayes(fit2)
>   > topTableF(fit2, adjust="BH")"
>    
>   I use a very similar code to find differences between time effects and strain effects.
>   I have two strains (B and W) and two different time points (2 and 1 hours). I am trying to find genes which respond differently:
>   1. in B vs W, when comparing time points 1 and 2 (i.e. I compare BvsW in time point 1 to BvsW in time point 2)
>   2. in time in different strains (i.e. I compare B1vsB2 versus W1vsW2) 
>    
>   I am getting the same results, or they should be the same?
>   Could you comment on whether this is a right way to analyse time series?

It appears you want to test for the interaction between time and strain. 
  Both of your points (1 and 2 above) are essentially identical. You 
want to know what genes react differently over time in the two strains 
(and worded differently, but meaning the same thing - what genes react 
differently between the strains at different times). These two things 
are verbo-algebraically the same (yup, new word. My next move is to 
start a new Wikipedia entry describing exactly what it means ;-D)

>    
>   My code is below.
>    
>    
>   Thank you,
>   Lev.
>    
>   > temp<-rma(data)
>   > targets <- readTargets("Targets.txt")
> 
>>lev <- c("W.1","B.1","W.2","B.2")
>>f <- factor(targets$Target, levels=lev)
>>design <- model.matrix(~0+f)
>>colnames(design) <- lev
>>fit <- lmFit(temp, design)
>>cont.dif <- makeContrasts(Diff.Time=(B.2-W.2)-(B.1-W.1), Diff.Strain=(B.2-B.1)-(W.2-W.1), levels=design)

As the statements above are verbo-algebraically the same, these two 
terms are algebraically the same.

(B.2-W.2)-(B.1-W.1) = (B.2-B.1)-(W.2-W.1)

Hence you should get the same results from each contrast. And yes, this 
contrast does give you the interaction.

Best,

Jim


>>fit2 <- contrasts.fit(fit, cont.dif)
>>fit2 <- eBayes(fit2)
> 
>    
>   I am getting the same results for both coefficients:
> 
>>results<-decideTests(fit2, method="separate", p=0.01, lfc=1)
>>summary(results)
> 
>    Diff.Time Diff.Strain
> -1        61          61
> 0      44951       44951
> 1         89          89
>    
>   My target file is:
>   FileName         Target
>   3517    W.1
>   3518    W.1
>   3519    W.1
>   3520    W.1
>   3521    W.1
>   3522    B.1
>   3524    B.1
>   3525    B.1
>   3526    B.1
>   3527    W.2
>   3528    W.2
>   3529    W.2
>   3530    W.2
>   3531    B.2
>   3532    B.2
>   3533    B.2
>   3534    B.2
>    
> 
>        
> ---------------------------------
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-- 
James W. MacDonald, M.S.
Biostatistician
Affymetrix and cDNA Microarray Core
University of Michigan Cancer Center
1500 E. Medical Center Drive
7410 CCGC
Ann Arbor MI 48109
734-647-5623


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