Trouble with the bluescript vector

[DUSAN at physik.uni-bielefeld.dbp.de]

IO00865 at MAINE says:

>I have been trying to use Stratagene's pBluescript II SK (+-) vector for some
>subcloning. So far I have only sequenced one clone, but part of the polylinker
>region and part of the insert appear to have been scrambled. Has anyone else
>seen anything like that with this vector? Is it worth sequencing more clones
>in search of a good one? Any comments would be appreciated.
>
>                                                     Thanks,
>                                                     Ethan Strauss
>                                                     Io00865 at maine.bitnet

I have got similar results using the Klenow fragment after ExoIII/ExoVII
treatment of an insert (in order to introduce nesting deletions). However,
this was only true for one sequenced clone out of four or five, I think.
The deletions (I think I missed about 10 bases) are probably due to the still
abundant but low exonuclease activity of the Klenow fragment.


regards,

Dusan Zivadinovic
Department of Genetics
University of Bielefeld
W 4800 Bielefeld
Germany

email: dusan at physik.uni-bielefeld.dbp.de