PCR products generated from unpurified Salmonella DNA are degraded by thermostable nuclease activity.
Oligonucleotide primers designed from repetitive extragenic palindromic (REP) sequences were used to PCR-amplify Salmonella DNA. Unpurified template DNA, present in crude cell extracts, yielded an essentially identical banding pattern to that arising from the use of purified DNA. However, the PCR product derived from the crude preparations did not survive storage at 4 degrees C. This post-PCR DNA degradation, attributed to endogenous Salmonella nucleases, was inhibited by the addition of EDTA, or storage at -20 degrees C.
Gibson
J R
JR
AFRC Institute of Food Research, Norwich Laboratory, Norwich Research Park, Colney, UK.
McKee
R A
RA
D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics / Q000378:metabolism*; D003851:Deoxyribonucleases / Q000378:metabolism*; D004795:Enzyme Stability; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012475:Salmonella / Q000235:genetics*
D004269:DNA, Bacterial; D003851:Deoxyribonucleases