help needed with southern

martin LEACH leach at
Fri Aug 4 11:32:37 EST 1995

HOw are you doing the a bottle or a bag...

amount of prehyb doesnt really matter....for me..

for the hyb i always keep it 1million cpm per ml of fluid (go up to 5 mill/ml)

in bottle hyb...large bottles...about 20ml...small bottles about 10ml

...letting us know what method you are using will help..


PAUL CANNNISTRARO (pcannist at wrote:
:    I am working with genomic DNA (10ug per lane of gel) and I am somewhat at a
:  loss in figuring out the appropriate amount of hybridization fluid to use.
:  Miniatis recommends .2ml of pre-hyb per sq. cm of nylon membrane, which comes
:  out to 34ml for 14x12cm. However, the amount of hyb solution is not specified,
:  but one  could infer that it is the same, since it is suggested to replace the
:  pre-hyb with an identical solution. The controversy arises when I take into
:  account that Maniatis recommends 10-20ng/ ml of labeled probe during 
:  hybridization and 100ng of probe altogether. Thus, I should have 5-10ml of
:  hybridization solution?

.....          Martin Leach                Email:leach at 
   _|____      Dept. of Pharmacology       Phone: (617) 638-5323        
   / o  /      Boston Univ. School of Med. Fax:   (617) 638-4329         
 _/  |-/__==/  80 E. Concord St. (L603)
(BULLDOZER) \_ Boston MA 02118            "Not the old underpants 
               USA                                on your

My homepage:
Biotech companies on the net:

More information about the Methods mailing list