Question: Anova-like in Limma
0
gravatar for ivan.vechetti
12 days ago by
ivan.vechetti0 wrote:

Hello guys,

I currently have the following design: (Knockout=TAm and WT-Veh) in a time course (0,3,5 and 7 days)

> design
   fTam.0d fTam.1d fTam.3d fTam.5d fTam.7d fVeh.0d fVeh.1d fVeh.3d fVeh.5d fVeh.7d
1        1       0       0       0       0       0       0       0       0       0
2        1       0       0       0       0       0       0       0       0       0
3        0       1       0       0       0       0       0       0       0       0
4        0       1       0       0       0       0       0       0       0       0
5        0       0       1       0       0       0       0       0       0       0
6        0       0       1       0       0       0       0       0       0       0
7        0       0       0       1       0       0       0       0       0       0
8        0       0       0       1       0       0       0       0       0       0
9        0       0       0       0       1       0       0       0       0       0
10       0       0       0       0       1       0       0       0       0       0
11       0       0       0       0       0       1       0       0       0       0
12       0       0       0       0       0       1       0       0       0       0
13       0       0       0       0       0       0       1       0       0       0
14       0       0       0       0       0       0       1       0       0       0
15       0       0       0       0       0       0       0       1       0       0
16       0       0       0       0       0       0       0       1       0       0
17       0       0       0       0       0       0       0       0       1       0
18       0       0       0       0       0       0       0       0       1       0
19       0       0       0       0       0       0       0       0       0       1
20       0       0       0       0       0       0       0       0       0       1

I want to compare within the genotype as well as time-course. I've been following the Limma section 9.6.1 (Time course with replicates) and my strategy looks like:

 > colnames(design)
 [1] "Tam.0d" "Tam.1d" "Tam.3d" "Tam.5d" "Tam.7d" "Veh.0d" "Veh.1d" "Veh.3d" "Veh.5d" "Veh.7d"

> fit<- lmFit(dat, design)

> cont.Tam<- makeContrasts("Tam.1d-Tam.0d", "Tam.3d-Tam.0d", "Tam.5d-Tam.0d", "Tam.7d-Tam.0d", levels = design)

> fit2<- contrasts.fit(fit, cont.Tam)

> fit2<- eBayes(fit2)

I would do the same for the WT as in the manual. My question is, am I doing this right, to have an Anova-like test? If so, how can I get the differences? Based on I saw it would be:

>topTable(fit2, coef = 1, number=Inf, adjust.method = "BH")

I just want to make sure if I'm doing the proper design and comparisons and how to get what is actually changing.

limma • 62 views
ADD COMMENTlink modified 12 days ago by Gordon Smyth37k • written 12 days ago by ivan.vechetti0
Answer: Anova-like in Limma
1
gravatar for Gordon Smyth
12 days ago by
Gordon Smyth37k
Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia
Gordon Smyth37k wrote:

Your analysis seems fine from the code you've shown. So far you have obtained a list of differentially expressed genes for KO at 1 day vs KO at 0 days. You can perform any other comparison you want in the same way.

ADD COMMENTlink written 12 days ago by Gordon Smyth37k

Thank you for the input.

ADD REPLYlink written 11 days ago by ivan.vechetti0
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