Help! Plaques don't come up.

Pascal Mertens Pascal.Mertens at fundp.ac.be
Fri Jun 6 01:58:50 EST 1997


In article (Dans l'article) <338D4089.6DD5 at qub.ac.uk>,
a.wallace at qub.ac.uk.see.signature wrote (écrivait) :
 
> I would also be interested in receiving any advice or information about
> this problem, since I am having similar difficulties at the moment using
> XL1-Blue transfected with M13 and growing on ordinary LB agar.



I recently screened XL1-Blue containing a phagemid and superinfected with
M13KO7.
Plates contained H plate media (tryptone 10gr/L, NaCl 8gr/L, Agar 15gr/L).
Top was H top agar: (tryptone 10gr/L, NaCl 8gr/L, Agar 8gr/L) and was
maintained at 45°C.
For plating: 5ml Htop agar, O.5ml fresh XL1-Blue, 0.1ml infected XL1-Blue.
Note that this was used for an immunological screening.
Membrane was set on the plate after 5 hours; after removal, the plaques
were nice and H top agar was not disturbed.
2nd note: H plates have to be well dried to prevent condensation on the H
top and subsequent mixing of the plaques!!

Hope this helps.
(I've not much experience in lambda phage libraries screening, and I guess
this protocol is not good for this last purpose).

Pascal Mertens

-- 
Pascal Mertens
Laboratoire d'Immunologie et Microbiologie
URBM-FUNDP
61 Rue de Bruxelles
5000 Namur, BELGIUM
Phone: 32 81 724438
Fax: 32 81 724420



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