BIIIIG problem with top-agarose (fwd)

[Neil Harris]

---------- Forwarded message ----------
>Date: 4 Nov 1995 11:32:23 GMT
>From: Hilde Nilsen <hilden at unigen.unit.no>
>To: "bionet.molbio.methds-reagnts mail newsgroup" <bionet-news at dl.ac.uk>
>Subject: BIIIIG problem with top-agarose

>Hallo reader

>I am screening a genomic lambda libary. I am using host cells without 
>selection.
>The problem is that as I am transferring the plaques to Hybond N+ 
>membranes, 
>the top-agarose adheres to the membrane thereby ruining everything.
>I suspect this to happen because of moisture between the top-agarose and 
>the medium.

>So, what can I do to prevent this? I am cooling the medium to 50 deg. 
>celsius before
>preparing the plates so too hot medium should not be the problem.

>Please  help me!

>My e-mail address: hilde.nilsen at unigen.unit.no



Dear Hilde,
	Try putting your plates in the fridge for at least two hours (or 
overnight if more convenient) prior to performing lifts on the lysed 
plaques. This should prevent or at least lessen the amount of top 
agarose sticking to your filters. It seems to be that the warmer the 
agarose is the more easily it will adhere to filters, hence it's not a 
very good idea to perform lifts straight after removal from the incubator.
	Hope this helps!

Neil Harris,
St. Andrews University.