[CLONING] denatured fragments before ligation

Ferdinando Pucci brotizolam at libero.it
Wed Jul 21 11:16:22 EST 2004


Ok thank you, i'll tell something more. It's a step to create a 
lentiviral vector from dTNS9-dMluI-dPmeI-dSmaI with inserts from 
pSLDT1180. Genes involved are thalassemia mutants of beta globin (i 
dunno how this help whatever :-). RE used: NcoI and SwaI. Vector 
excised (out normal beta globin) and ligated with betamutans.

I've yet done this before with another clone, but i've used an amicon 
ultrafree gelneb to elute from agarose, but with 10% yeald. This time i 
wanted to use old grandmama method ;-) and as foreseen i got better 
yealds! But at the end... :-/

Is it enough? :-)
Thanks again

Il giorno 21/lug/04, alle 17:41, Wolfgang Schechinger ha scritto:

> </kernel panic> ;-)
>
> Dear Fernando,
>
> would you give us a short description of what you actually plan to do, 
> please!
>
> (source of DNA, RE-enzymes used, strategy of cloning or whatever you 
> want
> to do)
>
> please be as specific as possible.
>
> Best regards,
>
> Wolfgang

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