un691cs at genius.embnet.dkfz-heidelberg.de
Wed Dec 11 13:36:39 EST 1996
In article <32AE92E5.7488 at glaxowellcome.com>, kucera~gt at glaxowellcome.com says...
>> I want to know if anyone has a formula similar to Clontech's expresshyb
>> for nucleic acid hybridizations?
>I have used the following QuickHyb solution (BRL's recipe)
>for a long time with great success on colony blots and
>southerns. I typically do a 1 hr. prehyb followed by 2 hr.
>hyb. for colony blots. Works great!
>I hyb southerns a little longer, >4 hrs.
>QuickHyb Recipe: I usually make up about 100mls then aliquot
>into 10-12 ml tubes and freeze.
>6x SSC ; 5x Denhardt's Solution ;0.5% SDS ;100ug/ml SS DNA
Sorry to correct you, but colonies contain an afwul lot of target
DNA, and the southerns that you mention are presumably NOT
genomic southerns, but PCR or restriction frags ?
Depending on WHAT you want to detect, such short hyb times are
not extraordinary.... However, in the buffer that you describe
a genomic southern would still take at least 16 h....
BTW, leave out the Denhardt's and use 7% SDS: easier
and less background.
So anybody know the recipe of a true quick buffer ?
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