Is Too Much Phosphatase Bad?

Michael Cooley cooley at mbcl.rutgers.edu
Wed May 21 06:35:56 EST 1997


BMB says that the problem of too much phosphatase (CIP) is that the
enzyme is difficult to get rid of. It binds to the ends of the DNA and
limits ligation. Their method to get rid of the phosphatase worked for
me. That is to add 0.1 vol 0.2 M EGTA pH8, heat to 65 oC and extract
with phenol/chloroform. The EGTA kelates zinc which is part of the
enzyme and helps to release it from the DNA. I found this method
effective even at 10X higher than recomended CIP concentrations
-- 
Dr. Michael B Cooley
Waksman Institute
Rutgers University
Piscataway, NJ 08855

908-445-3802
FAX 908-445-5735
Email cooley at mbcl.rutgers.edu



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