End labelling a primer

January Weiner jweiner1 at aixterm1.urz.uni-heidelberg.de
Fri Mar 20 14:34:55 EST 1998


[John]:
> Does anyone have a protocol for end labelling a PCR primer?  Thanks in
> advance.
> JPF
	3' or 5'? Radioactive, non-radioactive? Quick and dirty, or long
and cumbersome? As for 5' radioactive q&d we use
	(vol. 10 ul)
	30 uCi [gamma-32P]dATP (~3ul)
	5 pmol primer
	1 x PNK buffer (1 ul 10 x)
	10 U Polynucleotide Kinase (PNK)
	TE ad 10 ul

	incubate @ 37 deg. C. for 40', add 100 ul stop mix (Phenol, EDTA),
purify with Sephadex G-25 column / spin column.

	Regards,
			January

-- 
----)-\//-///-----------------------------------January-Weiner-3-------
His ideas of first-aid stopped short of squirting soda water.



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