Question: Some problems of creat DESeqDataSetFromMatrix step by DESeq2
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8 months ago by
110651827160 wrote:

For this step:

dds < - DESeqDataSetFromMatrix(countData = counts,
                                  colData = coldata,
                                  design = ~condition)

I read a lot of examples on the website, but these examples seems for majority cases, here are three parts I couldn't make sure:

  1. countData = counts
    Generally, part of counts matrix like the following, conl1 conl2 are two samples,

                  conl1 conl2 rep1 rep2    
    ENSMUSGid1     0     0    3    4    
    ENSMUSGid2     0     1    0    0    
    ENSMUSGid3     0     1   10   12

    but for my data, even as for conl1, here are three experimental replications, so, it means, for colnames of my raw countData:
    conl1_1, conl1_2, conl1_3, conl2_1, conl2_2.... I plan just seem all of them(conl1_1, conl1_2, conl1_3) as conl. Is it okay?

  2. colData = coldata
    Can this coldata contains all potential variables I want to study, including cell type column, batch column, gender column, age column? And let design = ~condition1 + condition2 +condition3...(Order according to proportion from PCA)? Or study each variable one by one?

  3. design = ~condition
    Actually my colnames of raw countData also contains batch information, generally, people use design = ~condition, can I use design = ~batch + condition (I gave one column of batch for colData = coldata)? Or run a design = ~condition DEseq2 result then design = ~batch DEseq2 result? Or I should check batch effects exist firstly, then corrected, then do design = ~condition?

Thanks for your attention in advance, sorry for so many confused... Any ideas of each part will be very grateful!!!

rna-seq R • 228 views
ADD COMMENTlink modified 8 months ago by lakhujanivijay4.3k • written 8 months ago by 110651827160
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