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qPCR and siRNA - (Jul/19/2005 )

I am a beginner with qPCR and I would like to check the effect of an siRNA with this technique. But the mRNA I am looking at seems to be at low copies. I start having an amplicon at around 30 cycles. It makes it difficult to see the effect of the siRNA as it is at the end of the run... How could I improve this experiment? Thanks for any advice...


I have the same problem, my gene of interest comes up after 45 cycles which is too late.

I checked some techniques like making aRNA described by Eberwine (on sale at Ambion) but this seems very labourous and expensive. I hope somebody knows an easier and more economical way?


well, but it should be OK?? is not it? If you knock-down your gene and after that it is hardly detectable, so you could be happy or not? i read somwhere that positive detection after 35 cycle could be considered as negative result.