dephosphorylation by SAP
hroychow at NMSU.EDU
Thu Jul 11 14:44:32 EST 1996
I had been using SAP lately. I use it the same way as I would CIP,
ie. adding it directly to the plasmid following restriction, and
incubating it at 37 C for an hour. In case of CIP this works fine. My
question to those who are using SAP from USB is 'does one need to use the
USB-supplied SAP buffer for the reaction, or does the regular RE buffer
(mine is a Tris-acetate, pH 7.6 with Mg and K-OAc) work for SAP
activity?' The buffer supplied is a Tris pH8.8 with Mg.
Obviously, I am not happy with the degree of dephosphorylation and
was wondering if our storage (-20 C) of the enzyme had anything to do with
it. We have not aliquoted it, and folks do use it from the same stock tube
a few times a week.
Hiranya S. Roychowdhury
Plant Genetic Engineering Lab.
Box 3GL, NM State Univ.
Las Cruces, NM 88003
Phone: (505) 646-5785
hroychow at nmsu.edu
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