Transformation question?

Martin Luetzelberger R333 5784 luetzel at alpha.bio.nat.tu-bs.de
Fri Jun 6 07:08:06 EST 1997


L M wrote:
> 
> Dear colleagues,
> 
> I'm doing in vitro mutagenesis by Kunkle's method. After I synthesized 2nd
> strand and ligated, then tranform host TG1. surprizingly I got no
> colonies, but the control plate(normal ds DNA) has a lot of colonies
> (>1,000/per ng).
> 
> The synthesis is completed by running a 0.7% agarose gel in the presence
> of 0.5ug/mL EtBr). My synthesized DNA band has the same migration as the
> supercoiled DNA.
> 
> The ssDNA must contain Uracil, because the titer of my uracil-containning
> phagemid in TG1 and CJ236 is very different(>10,000 folds).
> 
> Could someone tell me the possible reason?
> 
> Thank you in advance
> 
> Richard

Richard,

take an other strain e.g. XL1-Blue or a strain with an endA1 marker.
This should solve your problem.

Martin



More information about the Methods mailing list