Blunt-end troubles

Pete pkursula at cc.oulu.fi
Wed Mar 5 05:51:03 EST 1997


On Tue, 4 Mar 1997, T.J. Young wrote:

> Hi,
> I've got a bit of a problem, I'm trying to subclone a DNA sequence from one 
> vector into another.  Unfortunatly, the MCS are not compatible so I'm using 
> blunt-ended ligation.  Now this is a bugger at the best of times but every 
> time I get some positive colonies, on inspection, I find that the sequence 
> has gone in the wrong way round!!  Now am I just the Worlds unluckiest 
> molecular biology student or could there be some kind of underlying 
> phenomenon that's driving my ligation in this direction?
> Cheers,
> Tim ;-)
> 
Yes, it is a general phenomenon: your insert always goes in in the wrong 
orientation, if it is possible. 

;-) At least that is my experience.




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