Preparation of DNA extracted from environmental water samples for PCR amplification

Sephadex G-200 spun columns have been used to purify DNA extracted from aquatic samples. Nucleic acid recovery using a previously-described protocol was only 10 to 15%. We optimized this method by employing a high salt (0.2 M NaCl) TE buffer (pH 8.0) and four slow-speed centrifugation steps (130xg) in a swing-out centrifuge. DNA recovery improved to approximately 75%. Purified DNA was a suitable substrate for PCR amplification. Following PCR of DNA extracted from water samples amended with various concentrations of enterotoxigenic Escherichia coli (ETEC), the ETEC heat-labile enterotoxin (LT) was detected in samples treated with the Sephadex G-200 spun columns and not in untreated samples. This method has the advantages of being inexpensive, simple and rapid. (C) 1998 Elsevier Science B.V.