Question: Finding DEGs using RNA-seq data
gravatar for noorpratap.singh
3.1 years ago by
noorpratap.singh280 wrote:

Hey, it might seem a basic question but I wanted to find degs using RNA-seq data. First thing for finding degs should I take only tumor and tumor adjacent normal or should I compare the entire tumor samples even though for the patient I might not have normal samples against the few normal samples.

Further before finding DEGs the normalisation which we have to do has to applied on the entire samples and not only on those belonging to the same pateint?

At the moment I have a count matrix but as such if I look at total ensembl ids(row names) they are more than 60000. I understand they might contain noncoding regions so I map them to entrez genes but even after that I have some 24000 ids. Any further filtering which I need to apply.

Thanks in advance.

rna-seq degs • 1.1k views
ADD COMMENTlink modified 3.1 years ago • written 3.1 years ago by noorpratap.singh280
gravatar for Devon Ryan
3.1 years ago by
Devon Ryan91k
Freiburg, Germany
Devon Ryan91k wrote:
  1. You can exclude any samples without matching controls.
  2. You should be using a package from Bioconductor like DESeq2 or edgeR. This will have methods for normalization, just read the appropriate vignette.
  3. The genes should be rows, not columns, so transpose your matrix. Do not filter further unless it's by something like mean expression level (this is again covered in the aforementioned vignettes).
ADD COMMENTlink written 3.1 years ago by Devon Ryan91k

Thanks, 1)It was typing error genes were rows. 2)For normalisation do I need to exclude the other samples.

ADD REPLYlink written 3.1 years ago by noorpratap.singh280

You can keep all the samples in for normalization.

ADD REPLYlink written 3.1 years ago by Devon Ryan91k
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