Trimming 5' protrusive ends?

Cocea cocea at bisance.citi2.fr
Fri Sep 29 07:19:02 EST 1995


Hi- I have tried to blunt a XbaI (5' protrusive) DNA fragment end with the Mung
Bean Nuclease and my ligation did not work. I have been told that this enzyme
doesn't always work very well, so I might use the S1 nuclease instead. However,
I have only used S1 in a ExoIII/S1/Klenow protocol to create deletion 
mutants, so the Klenow enzyme was there to fill in any remaining protrusive
ends. 

What I would greatly appreciate is any advice on HOW to trim a 5' protrusive 
end so that it becomes blunt and I can ligate it to another blunt fragment.
It happens that I can't use the filling in approach this time. Any suggestion?

Thank you very much.

Laurentiu





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