Colony Hybridization



	1.	Using a black pen, mark the filter (Amersham's Hybond-NTM, 

82 mm dia.) to identify the plate.



	2.	Lay the filter on the agar surface to make a replica of 

the colonies.  Leave for 1-5 minutes.



	3.	Using a needle, pierce the filter and agar in an asymmetrical 

pattern to mark the position of the filter on the plate.



	4.	Carefully lift the filter off the plate and place colony-side 

up on a piece of Whatman 3MM paper.  Return the agar plate to the 37o C 

incubator to allow the colonies to grow.



	5.	To lyse the colonies and fix the DNA to the membrane, autoclave 

the filter(s) @ 212o C for 1 minute on the liquid cycle.  (Note:  The 

autoclave on the 6th floor of the Taub Building works the fastest, with the 

entire cycle taking about 8 minutes.)  Remember to reset the temperature 

on the autoclave to 250o C after use.



	6.	The filter(s) may be prehybridized and hybridized according 

to the standard Southern protocol.