kits and kits and kits and mutagenesis

Basavaraju Shankarappa bsh at MED.PITT.EDU
Sun Nov 15 13:10:09 EST 1992

>From: wsun at (Fiberman)
>I am trying to do site-directed mutagenesis using the Bio-rad
<kit.  I am using CJ236 cells and my clone in the Bluescript
>plasmid to make SS DNA with VCM13 helper phage.  Following the
>protocol, I do a negative control where I add everything except
> 	the mutagenic primer.  However, when I transform XL1-blue cells
>with this negative control DNA, I get colonies on the plate.  In
>addition, when I screened the colonies from my positive plate, I
>don't get any mutants, only wild type.  Can anyone out there
>have had this problem and could offer some


One suggestion regarding the above type of questions.  I think it will
be best for both people who read it and for those who post it, if you 
can please explain the method used in any of the above kits.  As you
all know many companies make more than one kind of kit and there are 
so many kits out there it is not possible for anyone to keep track of  
all types of kits.  Although it is very easy for us to think that the
procedure or the kit we are using is the best in the whole wide world
it is highly likely that many people never heard of it.  Also, we may
not have the time or patience to go the company catalog and see what 
the procedure involves.  If you explain the method, may be somebody
who used the method but not the kit will be able to help you.
	I think it is this presumption in kit users that infuriate the 
people of Jim Graham types.  I can very easily understand their 
frustration at having to work with pro-kit-robotic people.  As a 
cautious supportor of kits, this is just my opinion.  We need both
Jim Graham and Steve Mosley.  
Raj Shankarappa
bsh at

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