Can do reverse transcription by gene-specific primers? - (Feb/25/2008 )
Hi there,
It is ok to reverse transcribe RNA using gene-specific primers then using the cDNA to do real-time? For gene expression study. I personally think its ok because you do real time to detect the original amount of cDNA (or mRNA) of your gene. If so then the real time detects the gene-specific cDNA present. What's your opinion? WOuld like to know what the rest of the world thinks. Thanks and good day.
Chris
Yes it's ok!!! I've used this methods for a particular application, in which I'd no other choice.
Anyway you can do it for you normal expression study! 
If it was a normal RT PCR I would say yes, of course. But since you are doing qPCR I wander if it wouldn't affect your PCR efficiency since the concetration of the primers won't be optimal, especially for the one that you have used to reverse transcribe...
It is ok to reverse transcribe RNA using gene-specific primers then using the cDNA to do real-time? For gene expression study. I personally think its ok because you do real time to detect the original amount of cDNA (or mRNA) of your gene. If so then the real time detects the gene-specific cDNA present. What's your opinion? WOuld like to know what the rest of the world thinks. Thanks and good day.
Chris
Usualy you use a reference (housekeeping) gene to normalise your target gene expression in quantitative PCR. This way you cannot use the same template for reference gene and target one. And if you do reverse transcribing with gene-specific primers for the reference gene as well, you're now facing maybe a different RT efficiency AND different qPCR efficiency instead of only the second one.
I would personally choose classic random and/or oligo-dT primers for generating cDNA if you want to amplify more than one gene (which you should anyway for expression studies).