MacConkeys instead of Xgal/IPTG selection?

wetsel_r at msdisk.wustl.edu wetsel_r at msdisk.wustl.edu
Sun Aug 29 08:04:35 EST 1993


In a previous article, rrumpf at magnus.acs.ohio-state.edu (Robert Rumpf) wrote:
>I've heard that the alpha-complementation-based color selection using 
>lacZ-phagemids, lacZ deletion host cells, and Xgal/IPTG plates can also be done
>using the same phagemids/host cells but plated onto MacConkey's agar.  Is this 
>true, how is it done, and what's the mechanism for color selection?  I 
>appreciate any help you can give...

Hi Bob:  I'd strongly suggest that you look up
Jennings and Beacham in Biotechniques vol 7:1082 (1989).
Once our lab lacthed onto this method we rarely plate our ligations on 
anything else unless dictated by a particular vector that doesn't have the  
Betal-gal.  What is critical is that the bug of choice have the lac 
deletion.  HB101's don't work, but Sure's, JM109, TOP10F' and others work 
just fine.  At the time of plating you just put in 10 ul of 100mM ITPG and 
voila!  You have red/white color selection.  One thing though, over time 
your white (recombinant) colonies will turn a slight shade of pink making 
colonly selection after about 24-48 hours very difficult.  So if you do 
your ligations one day and will need to pick on another (for whatever 
reason), then one must mark which ones to take or not to take.

Hope this helps,
David

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