standard curve preparation - (Jun/29/2007 )

Is it possible to construct the standard curve by using cDNA.

first of all I will determine the its concentration by Spectrophotometricly and than cDNA will be diluted by using genome copy number. and RT-PCR will be performed. It is possible?

I don't think that will work.

the genome copy number/size are not likely to be proportional to the amount of message you had...and not all cDNAs act the same in the RT reaction.

do you have your DNA present on a plasmid? you can use that. if not, you can use purified PCR product, but I think it's a bit easier and more of a sure thing to clone your DNA into a plasmid and use the plasmid as a template

good luck!

ps, have you read User Bulletin #2, on ABIs website? it is a bit dense, but it contains what you need to get started