[BioC] LIMMA: data with technical replicate/familial relationship/biological replicate

[Steve Lianoglou]

Hi Qin,

I saw this question a few days ago here, and since I don't think
anyone tried to answer, I'll try to get the ball rolling with a
question of my own:

> say I have the following samples:
>
> f1_A1
> f1_A2
> f1_B1
> f1_B2
> f2_C1
> f2_C2
> f2_D1
> f2_D2
>
> Here
> f1 and f2 are two families
> A1 and A2 are technical replicate. same for B1 and B2,C1 and C2, D1 and D2.
> A,B,C,D are biological replicate, say A,C are affected while B,D are unaffected.
> Noted some families have more than 1 affected or unaffected samples.
>
> How does LIMMA take the two-level (technical replicate and familial relationship)
> dependence into account?

How would you expect limma (or anything) to take "familial
relationship" into account? Why do you want to?

I'm not sure what the right answer is, but I'd just recommend doing
some exploratory analysis -- perhaps it will help you find a
reasonable thig to do:

1. Look at each "family" separately -- f1_(A1,A2) vs. f1_(B1, B2) and
then f2_(C1,C2) vs. f2_(D1,D2), where you just use the expts in (...)
as technical replicates. Do the differentially expressed genes between
cases vs. normals differ between families?

2. What if you try a similar approach but combine families as
biological replicates and do cases vs. controls.

3. Is there any differential expression between "normals" in f1 vs.
f2? How about the cases in f1 vs. f2.

4. Which samples cluster together when you do a heatmap?

To help broaden your horizons and give you another perspective,
perhaps you can take a look at some of the GWAS papers from the hapmap
project. It's not really my field of expertise, but I know (imagine)
they need to deal with population structure/stratification issues,
which I guess you think is happening between your families (and why
you can't just use them as biological replicates?).

Of course you don't have anything close to the numbers, so everything
about your case is different, but there's some homework you can do (or
not) until someone gives you a more sound answer.

-- 
Steve Lianoglou
Graduate Student: Computational Systems Biology
 | Memorial Sloan-Kettering Cancer Center
 | Weill Medical College of Cornell University
Contact Info: http://cbio.mskcc.org/~lianos/contact