Optimization of metagenomic DNA extraction from activated sludge samples

Abstract

Metagenomic DNA extraction is essential for metagenomic technology. Therefore, optimization of a conventional total DNA extraction from activated sludge was investigated in detail in this study. Throughout two distinct orthogonal experiments, it was shown that the highest yield for metagenomic DNA could be obtained using TENP buffer, lysozyme of 1 mg ml-1 (1 h), protease K (200 μg ml-1), SDS (1%, 1 h). Furthermore, the quality of the differentially extracted DNA was subsequently assessed by the molecular fingerprint technology, such as denaturing gradient gel electrophoresis (DGGE) and ribosomal intergenic spacer analysis (RISA). The results indicated that the microbial diversity was dramatically different by different combined methods, and the DNA template quality for RISA was much better than that for polymerase chain reaction (PCR)-DGGE. This study provides detail process for metagenomic DNA extraction of activated sludge, and it would be useful for metagenomic DNA extraction of other environment samples. © 2009 Curtin University of Technology and John Wiley & Sons, Ltd.