Differencial expression analysis between groups determined by two variables

Question

[jeni]

Hi!

I am trying to perform a differencial expression analysis between two groups of patients. But I would like to group them taking into account two variables.

Imagine I have these data:

patient1  T1    6
patient2  T1    2
patient3  T4    2
patient4  T2    0
patient5  T3    1
patient6  T2    4
patient7  T3    5
patient8  T1    0

Would it be possible to group patients in two groups (for example: early and late) taking into account both variables? As an example: early patients could be those presenting T1 or T2 status referring to the first variable and a value shorter than 4 referring to the second variable. I wouldn't like to be subjective, so I would like to use some program which could perform this grouping.

On the other hand, would it be possible to create a design matrix with the two variables and then use limma to perform the differencial expression analysis taking into account both variables to group patients?

[swbarnes2]

Are you asking about software limitations, or the statistical soundness of doing what you propose?

[jeni]

About a method to do this with statistical soundness

[brunobsouzaa]

I would suggest you to use Rsubreads to create your matrix just inputting the corresponding samples for each group and then use Deseq2 and/or EdgeR for DE. But I think that you could use RSEM to generate your count matrix for using it with limma, that should be fine!