T4-pol PstI/Exo EcoRI/fill in

Ricky Boernke ricky_boernke at gmx.net
Tue Oct 30 03:18:16 EST 2001


Dear all,

I would like to remove all restriction sites but the HindIII site from
the pUC18
polylinker region. Therefore I would like to cut the vector with
PstI/EcoRI and
subsequently blunt the ends and religate. To obtain blunt ends I was
thinking of
using T4-pol since it should nibble back the PstI overhang and fill in
the EcoRI
site, actually within the same reaction. Well, I am always a bit
confused about the
exo and pol activities of T4-pol under different conditions so I am not
quite sure
whether this will work. Or will I have to do the blunting in seperate
reactions?
Any hints appreciated.

Ricky




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