cutting large circular DNA molecules

Bob bbx107 at excite.com
Tue Apr 19 23:31:55 EST 2005


On Tue, 19 Apr 2005 19:47:19 +0200, The Duck <theduck at earth.com>
wrote:

>Dear All,
>
>I was hoping that you could give me some advice to the following problem:
>
>I would need to cut large circular DNA (several hundreds of kilobases in
>size) exactly once (or at most only a few times). 

You hedged there in (). That makes a big difference. Cutting exactly
once -- without knowing anything -- sounds hard. Cutting "not much"...
Why not use a rare-cutting restriction enzyme? An enzyme with an
8-base recognition sequence cuts, on average, every 65 kb.

Look at the New England Biolabs catalog. Their discussion of megabase
mapping may help you.

bob


>I do not know the
>nucleotide sequence of the molecules and additionally they will not be the
>same. Currently I am trying a number of restriction enzymes, however, this
>is presently a workaround. What I would really need is a method that is
>cutting these large molecules only once.
>I was already thinking of using radioactivity, however, we cannot do this
>were I am working due to security constraints.
>I would very much appreciate any ideas you could give me.
>
>The Duck.




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