Choice of one-step or two-steps RT-PCR - (Jul/09/2008 )
I would like to perform RT-PCR. As you know, however, there are two types of RT-PCR, one-step and two-steps RT-PCRs. Could you tell me which RT-PCR you recommend and the reason ?.
It depends on your downstream applications. If you would like to analyse several genes, then you should consider two-step.
I prefer 2-step; my reasons:
- you need less RNA
- it's easier to optimize
- it's more reproducible
If you need to amplify very few targets, 1-step might be an option. Otherwise, I would not think twice and go for 2-step.
Thanks for the replies.
I would like to perform RT-PCR for the cloning of a single gene. Almost one-step RT-PCR kits use Taq DNA polymerase and are often used for the detection of gene expressions. I wonder if one-step can use for gene cloning.