topo ta cloning

ChenHA hzhen at freeuk.com
Thu Dec 30 14:27:10 EST 2004



"D.K." wrote:
> 
> On Wed, 29 Dec 2004 20:17:09 +0000, ChenHA <hzhen at freeuk.com> wrote:
> 
> >The reason is usually toxicity of the gene product.
> 
> I imagine that could be the case but it's not the explanation I like. The
> plasmids we are talking about are straight cloning vectors with no promoters,
> ribosome binding site, etc. The toxicity would have to be extreme - which is not
> the case of the same genes expressed later in pETs.

I haven't tried Topo TA cloning before (in fact, I always clone
directly into expression vector, and none of these expensive Topo
stuff), so I can't say what what might be the cause exactly in this
case.  I have found that, however, vectors that aren't supposed to
express a protein often do, albeit in tiny amount.  The cells may only
be growing somewhat slower, i.e. there might be the same number of
clones in both orientation but the colonies are much smaller or not
visible the next morning.


 
> DK



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