Bis. PCR not working. MSP is fine - (Aug/21/2006 )
I have a problem with my Bisulfite PCR amplification. Basically I can't amplify my bisulfite treated DNA.
I am using previously published primers.
I have done the following controls:
#1. I have used the DNA for MSP PCR and it works great!
#2 I have methylated a plasmid containing the region I am interested in with SSS1 and
did a bisulfite conversion of it. I then used my bisulfite primers on this construct and they amplified great.
P.S. This is a great control for people to incorporate into their work.
So, any suggestions for things to try or ideas why my amplification nof my bisulfite treated genomic DNA isn't working?
Can you post your PCR conditions for both MSP and BSP?
Usually, BSP requires more cycles than MSP. Nested PCR or two rounds of PCR using the same primer pair are generally required to amplify products for sequencing or cloning.