Detection of ss DNA intermediate

Stephen Billington billing at vetsci.microvet.arizona.edu
Mon Jul 21 13:03:21 EST 1997


Hi all

I am currently attempting to show that a plasmid replicates by rolling
circle replication by detecting single stranded intermediate. This is
usually done by Southern blot without denaturing the DNA prior to
transfer. However, everytime I try to do this I get double stranded DNA
transferring to the blot. I know this because my control which is just a
pUC based plasmid lights up in the Southern. Does anyone have any
experience with this sort of thing and can perhaps help me? This is
particularly frustrating as it seems to be a standard procedure and I
have a manuscript waiting for this one experiment.

sjb






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