Is it possible that I see strong repression of a gene by shRNA using Q-RT-PCR and immunoblot, but not on gene expression array (Human ST 1, affymetrix)?
The simple answer is yes. Two possibilities come to mind. First the probe set on the array for your favourite gene may not be working. The other possibility is that your array probset is recognizing the shRNA sequence used to KD your target. I have experienced both. I would avoid the ST arrays altogether, as they flatten data. I have had much better luck with the 133 arrays from Affy.
Best of luck,
Gavin
Dear Gavin,
thank you very much for the answer. I came to the similar conclusion with help of Karlheinz Holzmann. When he looked at specific transcription clusters, 3 of 6 (exon 2 and 3) were strongly downregulated in shRNA groups. Another 3 of exon 1 did not show any differences.
- Badges
- Science method