Help! - EMS mutagenesis

Brad Nicholson brad at corona.med.utah.edu
Mon Jul 10 13:08:49 EST 1995


In article <T.Barnett-0907951236160001 at 131.217.101.115>, 
T.Barnett at path.utas.edu.au (Tim Barnett) wrote:

> I am trying to mutate an endogenous alkaline phosphatase gene from
 >Aeromonas sobria and intend to perform subsequent TnphoA mutagenesis on
 >this strain.  
>deletia<
Hi Tim, 
I take it that you haven't found your mutant of choice yet.  First a couple
questions and then a suggestion.  
1) Are you sure that Aeromonas sobria has only one phosphatase?  If there 
are multiple phophatases that are active on LB/BCIP plates then the chances
of seeing a white is much lower.  
2) Have you done a killing curve with your mutagenesis yet?  This is a 
handy thing to do in order get a handle on how hard you are hitting your 
bugs. 

A realy good resource for your experiment is Jeffrey H. Miller's "A Short 
Course in Bacterial Genetics" 1992. Cold Spring Harbor Laboratory Press. 
ISBN 0-87969-349-5.  Specifically, Experiment 3, Mutagenesis with EMS.

Good Luck,
Brad

===========================================================================
Brad Nicholson              |"If it worked the first time, it wouldn't be 
Department of Pathology     |	research."...Brad Nicholson
University of Utah          |	Live from behind the Zion Curtian.
Salt Lake City, UT 84132    | 
brad at corona.med.utah.edu    |
or: (801)-581-4365          | My opinions are solely my own.
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