I am puzzled by discrepancies between DGE results obtained with SOLID and Q-RT-PCR confirmation in one experiment where we looked at the response of a cell line to one drug, after 24 hours: whereas we can confirm in all tested cases mRNA upregulation, we were unable to confirm downregulation: In each case (10/10) DGE indicates significant expression level (>500 reads) and clear downregulation, whereas no change was observed by Q-RT-PCR, on the same samples. PCR primers were in 3'Unt regions.
We made attempts trying to understand this discrepancy:
- using oligodT or random primers for Q-RT-PCR has no influence.
- Q-RT-PCR does not reveal downregulation at later time points (48h).
- biological repeats will be runned soon.
Did anyone already observed the same problem? Any possibility that DGE can go wrong?
We made attempts trying to understand this discrepancy:
- using oligodT or random primers for Q-RT-PCR has no influence.
- Q-RT-PCR does not reveal downregulation at later time points (48h).
- biological repeats will be runned soon.
Did anyone already observed the same problem? Any possibility that DGE can go wrong?
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