HELP!! transformation problems

Frank O. Fackelmayer fof1 at chclu.chemie.uni-konstanz.de
Wed Aug 5 10:36:32 EST 1998


Hi Ricky,
Yes, I agree that you have to grow fresh bacteria each time you want to do
transformations. But, well, the bugs grow on their own, and all you have to do
is to inoculate from a glycerol culture in the morning. Not too much work.

The main advantage of the method is that you don´t have to do more with your
bacteria than to spin down, resuspend, and add to DNA. No heat-shock, no
recovery period, no expensive special equipment. That´s why I prefer to use
it. Nothing even comes close with respect to hands-on time.

Gruss,
Frank


Frederik Boernke wrote:
> 
> Frank O. Fackelmayer wrote:
> >
> > For routine cloning, where maximum competence is not an issue, we always use
> > the TSS method of Chung et al.
> > It is very reliable once you got it to work.
> >
> > PS: Do not try to freeze the bacteria. They will go down in competence at
> > least a factor of 100. Well, that might still be ok for transformation of
> > supercoiled plasmids, but does not work for ligations any more.
> >
> 
> Hi!
> 
> When I get it right you always have to prepare fresh cells when planning
> a transformation, haven't you? So, what is the advantage compared to a
> protocol where you can freeze/store the competent cells for ages? In
> this
> case you prepare once a huge amount of bugs and from time to time you
> pull some out of the freezer.
> 
> Gruss
> Ricky
> 
> ******************************************************************
> 
> Frederik Boernke
> Research Group of  Molecular Plant Physiology
> Institute for Plant Genetics and Crop Plant Research (IPK)
> Corrensstr. 3
> 06466 Gatersleben
> Tel.  039482 -5 321
> Fax. 039482 -5 515
> http://www.ipk-gatersleben.de



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