real time PCR primer for methylation - (Mar/14/2005 )

I am trying to design primers for H19 by using primer express, because I want to use probe in my real time pcr. But the software doesn't give me anything. If I use sybergreen, the primers I am using right now have dimer, so it won't work. Anybody can give me some suggestions?

Hi zzylx,

I would like to ask one question, are you trying to design methylation specific PCR primers for a template that has been bisulfite treated? Or are you designing normal primers to the H19 gene, so you amplfiy off restriction digested genomic DNA?

I ask this because I don't think Primer express can design methylation specific primers. Check out http://www.protocol-online.org/forums/inde...?showtopic=4589 for specific primer designing tools.

I assume you would like to use the Taqman system, again I don't think Primer express can do this easily. However, you can "mock" bisulfite convert your sequence for primer express to pick appropriate primer and probe sets.

Good luck!

Nick

Hi,

Thanks for your suggestion. I am doing methylation PCR for bisulfite treated DNA. I input the bisulfite treated sequece to primer express, and got nothing.