Labelling PCR fragment for Northerns

Klaus Salger salger at wap1.zi.biologie.uni-muenchen.de
Tue Apr 22 11:35:15 EST 1997


jjm30 (jjm30 at dka.sm.ic.ac.uk.) wrote:
: Hi all,   I have a 500 bp fragment generated by RT-PCR, whats the best
: way (or easiest but good way) to radiolabel this for use as a probe in
: Northern blotting to detect a moderate abundance message.  I've seen
: the recent posts about labelling oligos but they seemed to be for
: smaller (100 bp) fragments.  Is random priming the best way to go??

Random priming should work fine. If you want to get a strand-specific probe
you may also use one of the PCR-Primers (the right one!) instead of the
random oligo mix. I've usually used Stratagene's Prime-It kit (exo-minus Klenow
, 9mer random  primers) which always gave high specific activity. 

Cheers
  Klaus


standard disclaimers apply

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Klaus Salger		phone : +49 (0)89 5902 	-502
Zoologisches Institut	FAX   :			-450
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