Plasmid for qPCR standard curve - (Dec/06/2006 )
Well, I have been handed this project of quantifying PCV2 (DNA virus) in field samples using real-time PCR. I have a few published protocols that seem to have a good amout of info, its just they they all seem to use this pIC2 plasmid for the standard curve. It is basically the PCV2 genome cloned into a pCR2.1 plasmid. I really know nothing about plasmids, is there somewhere I can buy the assembeled pIC2 plasmid, or do I need to somehow purify the genome and clone it into the pCR2.1 plasmid myself?
The pcr2.1 is a plasmid by invitrogen for cloning pcr products. One way to make the plasmid you discuss yourself it to clone the products of the pcr reaction using the primers for qPCR into the pcr2.1 topo vector. Once inside the vector, you can then calculate your standard curve. If you do not know how to do this write back.
I have a vague idea, any help you could provide would be appreciated.