5'-end labeling of 8mer oligo?
bhandary at wccf.mit.edu
Mon Nov 14 17:27:00 EST 1994
In article <9411120046.AA20356 at pollux.cc.umanitoba.ca>, pzhang at cc.UManitoba.CA (pzhang) writes...
>I posted my broblem a couple of days ago. Having got no reply, I'd like to try again.
>I want to radioactivly label 5'-end of a 8mer oligo. I am not sure how to separate labled oligo from the reaction (removal of ATP, ADP etc.) I want to use this oligo as a strand of an adapter. So, my second question is: If I use the labeled reaction dire
>Any ideas and suggestions are highly appreciated.
>pzhang at cc.umanitoba.ca
I would use either DEAE or PEI cellulose, developing in 0.75M KH2PO4. The ATP
will elute, and the oligo will stay at the origin, or move a bit off it. You
can cut out the spot with your labeled oligo, and elute with a high
concentration of some volitile salt buffer, either TEAB carbonate, or ammonium
An alternate approach is to use a Waters Sep-Pak column. Bind your sample at
low salt. Wash with 50mM TEAB carbonate to get rid of excess ATP, etc., and
then elute your oligo with 60% methanol. dry down, and dissolved in water or
the buffer of your choice.
More information about the Methods