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HELP WITH LIGATION OF PCR RESTRICTION FRAGMENTS

kanemats at angis.su.oz.au kanemats at angis.su.oz.au
Mon Aug 23 23:47:46 EST 1993


WANTED: PROTOCOLS FOR LIGATING RESTRICTED PCR FRAGMENTS

I would like any successful protocols available which
could allow the ligation of 3 PCR products (each of
approximately 200bp) after restriction digestion by PstI
(PCR products 1 & 2) or by BamHI (PCR product 2 & 3).
Detection of ligation product will be by PCR. I have
tried using approx. 1fg of each (Genecleaned) fragment
with 9 Weiss units of Promega ligase in commercial
buffer at 16oC overnight, and also 5-20ng of each (gel-
purified) fragment with 1.5 Weiss units for 1 hr at
18oC. Ligase has been used successfully by fellow
workers.

Francisca Springall
Kanematsu Laboratories
kanemats at morgan.angis.su.oz.au



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