[BioC] Do my Limma results look "normal"?

[Robert Gentleman]

Hi Paul,

   Please check the posting guide and provide us with the information 
requested there (like sessionInfo and the commands you ran). And I 
typically don't give any advice (excpet to follow the posting guide) to 
people who don't use signatures that identify them.

   Robrt


Paul Geeleher wrote:
> Hi,
> 
> This is the first time I've ever analyzed a microarray experiment
> using Limma (or anything else for that matter) and I was hoping that
> somebody could look at my results and tell me if they look normal.
> 
> The experiment is measuring differential expression between miRNAs of
> HER2+ and HER2- breast cancer tissue. There are 3 HER2+ arrays and 4
> HER2- arrays and each of the 399 miRNAs is replicated 4 times in each
> array.
> 
> TopTable() reveals the following miRNAs with a fold change above 1.5,
> which I thought was a reasonable cutoff:
> 
>                   ID     logFC         t      P.Value    adj.P.Val          B
> 273      hsa-miR-451 -4.645060 -8.226854 4.510441e-09 9.246404e-07 10.8484797
> 128      hsa-miR-205  3.551495  7.574564 2.370061e-08 3.239083e-06  9.2222865
> 13       hsa-miR-101 -2.310652 -6.569497 3.374177e-07 2.567796e-05  6.6146751
> 282      hsa-miR-486 -2.686910 -6.542808 3.626060e-07 2.567796e-05  6.5439656
> 55       hsa-miR-144 -2.890719 -5.889594 2.152998e-06 1.261042e-04  4.7952480
> 387      mmu-miR-463 -2.609257 -5.764143 3.042120e-06 1.559086e-04  4.4561920
> 388      mmu-miR-464 -2.080402 -5.696976 3.662006e-06 1.668247e-04  4.2743601
> 151      hsa-miR-223 -1.722956 -5.637290 4.318942e-06 1.770766e-04  4.1126276
> 51    hsa-miR-142-3p -3.262824 -5.397809 8.386312e-06 3.125807e-04  3.4626378
> 14   hsa-miR-101_MM1 -1.922710 -5.224075 1.358743e-05 4.175776e-04  2.9905370
> 159  hsa-miR-26b_MM2 -2.221853 -5.206724 1.425875e-05 4.175776e-04  2.9433849
> 236 hsa-miR-376a_MM1 -1.633555 -4.653220 6.637043e-05 1.700742e-03  1.4433277
> 266     hsa-miR-432*  1.512622  4.627293 7.131510e-05 1.719952e-03  1.3734422
> 168      hsa-miR-29b -1.954087 -4.198854 2.323860e-04 4.763912e-03  0.2280262
> 31  hsa-miR-126*_MM2 -1.537988 -3.209957 3.233842e-03 5.099520e-02 -2.2888897
> 52    hsa-miR-142-5p -1.881192 -2.831493 8.332384e-03 9.002153e-02 -3.1731794
> 
> 
> Another person is sanity testing this data using GeneSpring and they
> are getting much higher p-values compared to mine. They are also
> taking the step of excluding quite a few of the miRNAs from the
> experiment based on their standard deviation across the arrays of each
> group. Should I be doing this also or is this taken into account by
> the eBayes() function or lmFit()?
> 
> If you are interested the script I wrote to do the analysis is here:
> http://article.gmane.org/gmane.science.biology.informatics.conductor/18032/match=miRNA
> 
> Thanks for any advice,
> 
> -Paul.
> 
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-- 
Robert Gentleman, PhD
Program in Computational Biology
Division of Public Health Sciences
Fred Hutchinson Cancer Research Center
1100 Fairview Ave. N, M2-B876
PO Box 19024
Seattle, Washington 98109-1024
206-667-7700
rgentlem at fhcrc.org