Question: deseq2 analysis multiple factor and 4 different time points
gravatar for yusuf.zhc
13 months ago by
yusuf.zhc0 wrote:
I have multifactor data 
1. Participants are healthy population, having both males and females.
2. different excersice intensity for different legs. few participanrts have high intensity on left and low intensity in right, and few have high intensity of right and low in left.
3. samples were taken from both righ and left leg. 
4. samples were taken at 4 different time point
5. participants response to training, few participant show high muscle grwoth in high intensity some show high muscle growth in low intensity.

I am formulating this to normalize count table i got from htseq and rsem and trying to identify differentially expressed gene responsible for muscle growth and look if there is any biases based on leg, gender, excersice intensity. 

ddsHTSeq <- DESeqDataSetFromHTSeqCount(sampleTable = sampleTable, directory = ".", design= ~ condition)
dds <- DESeq(ddsHTSeq)
table_counts_normalized <- counts(dds, normalized=TRUE)
samples$Group <- factor(paste0(samples$participants,samples$response, samples$intensity, samples$time, samples$gender))
dds <- DESeqDataSetFromMatrix(countData = table_counts_normalized, colData=samples, design=formula(~Group))
dds <- DESeq(dds)
ADD COMMENTlink modified 13 months ago by swbarnes2340 • written 13 months ago by yusuf.zhc0
Answer: deseq2 analysis multiple factor and 4 different time points
gravatar for swbarnes2
13 months ago by
swbarnes2340 wrote:

You make dds from raw counts, not normalized.  I'm not even sure what happens if you try to make a dds object with non-integer counts as input.


That method of making a concatenated "group" column is useful for comparing subsets of samples to each other, while using the dispersion of the whole dataset, but I don't think that's what you want to do.  You probably do not want to compare one participant at one time with another participant at another time, but that's the kind of comparison you've set up there.  


You've also omitted the other key step in setting up DESeq; the call to results, which determines exactly what comparison you are making.





ADD COMMENTlink written 13 months ago by swbarnes2340

thank you for your reply.

I understand how to give concatenated subsets but i am not able to add time course into that. how should i compare then. 

ADD REPLYlink written 13 months ago by yusuf.zhc0
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