primer design problem - (Dec/16/2009 )
i am designing primer for SNP-RFLP and no restriction enzyme is available to cut the SNP site.
so, i make an artifical restriction site by inducing a primer mismatch in the primer, so that the pcr product can be cut by enzyme.
however, the mismatch i induced is near the SNP site, and after cutting by enzyme, the fragment is too short that cannot be visualized...
someone said i can add a poly(T) tail to the 5' end of the mismatch primer so that the fragment can be long enough.
anyone know whether it will work???
Yes, this will likely work. There is nothing magic about poly T -- an arbitrary sequence would also be fine (one that doesn't also get cut by your enzyme, of course).