Took a day off from coding…

Given my matrix.. these 2 blocks of code should have called different sets of results.. but the result of every (up/down) file is identical (now that I wrote them to excel and can look at them side by side…  the topTable for each data set is different for each contrast so printing that and can sort in Excel.

topTable(fit2, adjust="BH", n=20, coef=1, sort.by=”logFC”) #Bacteria - healthy
tops <- topTable(fit2, n=Inf) # get all genes
tops [which(tops$logFC > 0), ] [1:25,] # up reg top 25
write.table(tops, file="topsbacteria.txt")
tops [which(tops$logFC < 0), ] [1:25,] # down reg top 25
write.table(tops, file="lowsbacteria.txt")


topTable(fit2, adjust="BH", n=20, coef=2, sort.by=”logFC”) #phytoplasma - healthy
tops <- topTable(fit2, n=Inf ) # get all genes
tops[which(tops$logFC > 0), ] [1:25,] # up reg top 25
write.table(tops, file="topsphytoplasma.txt")
tops[which(tops$logFC < 0), ] [1:25,] # down reg top 25
write.table(tops, file="lowsphytoplasma.txt")

if I put a coef into the second line (for example)

> topbacteria <- topTable(fit2, adjust="BH", n=20 coef=1)
Error: unexpected symbol in "topsbacteria <- topTable(fit2, adjust="BH", n=20 coef"

But thus all my resultant up and down regulated tables are the same….

Whether you think you can or think you can't - Your're right...
Henry Ford

Cristina Paul

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From: seandavi@gmail.com<mailto:seandavi@gmail.com> [mailto:seandavi@gmail.com] On Behalf Of Sean Davis
Sent: Tuesday, May 01, 2012 1:30 PM
To: Paul, Cristina
Cc: bioconductor@r-project.org<mailto:bioconductor@r-project.org>
Subject: Re: [BioC] How do I find up and down regulated genes for each contrast in LIMMA?



On Tue, May 1, 2012 at 1:22 PM, Paul, Cristina <Cristina.Paul@ars.usda.gov<mailto:Cristina.Paul@ars.usda.gov>> wrote:
R.2.14 in RStudio all packages up to date.

Using the contrast.matrix I have been using lmfit etc from Limma, and have obtained a very informative Venn Diagram of our results (all trts minus healthy)...  I would like to find the top 25 (in one trt only 5 genes were unique to the pathogen)  up and down regulated genes... for each pathogen vs Healthy contrast. The code below gives me total across all the contrasts, or appears to.   Have googled and read most of day but  have found no code etc for doing this, or do I need to make a matrix for each comparison separately?     I tried topTableF in the tops code (given topTableF provided the contrast data) but result was NA's...

> design  #each trt 4 CEL file replicates no subtreatments
  Virus Bacteria Healthy phytoplasma
1    0        1       0     0
2    0        1       0     0
3    0        1       0     0
4    0        1       0     0
5    1        0       0     0
6    1        0       0     0
7    1        0       0     0
8    1        0       0     0
9    0        0       1     0
10   0        0       1     0
11   0        0       1     0
12   0        0       1     0
13   0        0       0     1
14   0        0       0     1
15   0        0       0     1
16   0        0       0     1


fit <- lmFit(eset, design)
contrast.matrix<-makeContrasts(Bacteria-Healthy, Phytoplasma-Healthy, Virus-Healthy, levels=design)
fit2 <- contrasts.fit(fit, contrast.matrix)
fit2 <- eBayes(fit2)
topTable(fit2, adjust="BH", n=20)
results<-decideTests(fit2)
vennDiagram(results)
topTableF(fit2, n=20)

tops <- topTable(fit2, n=Inf) # all on chip

You'll want to include the coef argument in the topTable call to specify which contrast you are examining.

Sean

tops[which(tops$logFC > 0), ] [1:25,] # up reg top 25
tops[which(tops$logFC < 0), ] [1:25,] # down reg top 25

thank you in advance for all help....

Whether you think you can or think you can't - Your're right...
Henry Ford

Cristina Paul






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