I ran qPCR arrays to test the expression of 48 genes in two samples. 10 genes showed significant difference between samples. The assay was very reproducible (nearly same results between replicates). I am wondering will I need to design specific qPCR assays for the 10 genes to confirm their expression before testing their protein expression?
In my opinion the decision depends on the number of replicates you tested with qPCR arrays; in my case I did the qPCR arrays (90 genes on the same sample) on three replicates. This is why I decided to design specific primers for a group of 15 genes, and to test their expresison on anotter 4 donors, to strenghten the data. It's up to you.
Allt he best for your experiments
Francesca Mariani
Many thanks Francesca, but I would kindly ask you; did the specific assays showed the same pattern of expression as in the arrays?
Thanks
M,ohamed
I'm still running the experiment, and I will let you know.
Francesca
In my opinion you need to replicate it in more samples. For all the genes where you find a change in expression, do a gene specific q PCR in more number of samples, say some 10-15 samples if possible. that way you can confirm it.
If your only concern is whether your should proceed to protein expression, then qPCR array results you got are probably justification enough. However, if you are going to rely purely on PCR results, then you need to do RT PCR for your 10 genes of interest in larger number of samples.
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