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Question: Design formula in DeSeq2 to control for a variable
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gravatar for Lakhan
9 days ago by
Lakhan0
University of Macau
Lakhan0 wrote:

 Hi all,

We have performed a RNAseq experiment to understand the differential gene expression upon repression of a gene. We have generated a strain in which the gene of interest is placed under doxycycline (DOX) repressible promoter (genotype: tet90). When the DOX is added to medium, gene gets repressed. DOX also has its own effect on the overall gene expression and hence we also have the RNAseq data for WT strain treated with DOX. WT strain grown on YPD medium is used as control. Please refer below for the complete design matrix for details:

genotype medium replicate
WT YPD 1
WT YPD 2
WT YPD 3
WT Dox 1
WT Dox 2
WT Dox 3
tet90 YPD 1
tet90 YPD 2
tet90 YPD 3
tet90 Dox 1
tet90 Dox 2
tet90 Dox 3

We would like to see the differential expression for gene repression condition vs normal condition (tet90_Dox vs tet90_YPD). However, we would also like to control for the DOX effect in this comparison. I think that this situation is similar to one of the Michael's reply in recent post on comparison (B-control_B) - (A-control_A)  C: help DESeq2 model design. I have tried exploring other similar posts on forums but still could not come up with a design formula. What can be the appropriate design formula to address this problem in DESeq2?

Thanks,

Lakhan.

ADD COMMENTlink modified 9 days ago by Michael Love18k • written 9 days ago by Lakhan0
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gravatar for Michael Love
9 days ago by
Michael Love18k
United States
Michael Love18k wrote:

If I read your question correctly, I think you want a design of ~genotype + medium + genotype:medium, and to test the interaction term to see if the ratio changes while controlling for the ratio in WT. Take a look at the Interaction section in the DESeq2 vignette, and the diagram there.

ADD COMMENTlink written 9 days ago by Michael Love18k

Thank you for your help Michael. Yes, your interpretation is right. I went through the DESeq2 vignette as well as the help page for results function. Example 2 in the results help page is similar with the case we have in our experiment design. As I understand, the interaction term genotypetet90.mediumDox will help us to identify the DEGs under geneX repression (by DOX treatment) in tet90 strain and at the same time control for the DOX effect in WT strain.

 

Thank you again.

ADD REPLYlink written 8 days ago by Lakhan0
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