Blotting of PFGE

Julia Lodge Lodgejm at bham.ac.uk
Mon Jul 22 16:57:25 EST 1996


I have a pulse field gel of bacterial chromosomal digests (500-1500kb 
approx) I want to transfer these to a nylon membrane for a Southern blot.
I have been advised that the old fashioned capillary blot works better 
than vacuum blotting. I have tried this with 2 X 15min washes in 0.2M 
HCl, the usual denaturation and neutralisation and an overnight transfer. 
On restaining my gel in the morning very little transfer seems to have 
taken place. Any advice? I wondered about electroblotting but can only 
find protocols for transferring up to 50kb fragments from agarose. Anyone 
tried electroblotting in this size range?

Julia Lodge



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