An effective method for extracting DNA from contaminated soil due to industrial waste.
Recently, labor accidents caused by toxic gas production such as H2S and CH4 have happened at some waste disposal sites and illegal dumping sites in Japan. The gas production has been known to be end products of bacterial respiration. In order to investigate a microbial flora in industrial waste contaminated soil by a DNA-based detection method, we developed a simple DNA extraction method. The method is based on lysis with sodium dodecyl sulfate (SDS) and a high concentration (4.0 mg/ml) of lysozyme at room temperature. DNA yields increased according to the concentration of lysozyme (0-2 mg/ml), although the cell lysis occurred at comparable efficiency independent of the lysozyme concentration. Interestingly, bovine serum albumin and insulin (4 mg/ml) also contributed high DNA yields. This suggests proteins play an important role at high concentration on a DNA extraction process, presumably by suppressing DNA degradation or adsorption. DNA yields from three contaminated soil samples at 0.3, 1.5 and 3 m in depth by using our method were significantly higher than those extracted by using a widely-used conventional method which did not contain lysozyme. A novel and simple DNA extraction method using high concentration of protein was established for a DNA-based detection method of microbial flora in soil.
Hinoue
Mitsuo
M
Department of Biological Functions and Engineering, Kyushu Institute of Technology Graduate School of Life Science and Systems Engineering, Wakamatsu-ku, Kitakyushu 808-0196, Japan.
Fukuda
Kazumasa
K
Wan
Yan
Y
Yamauchi
Kazuki
K
Ogawa
Hiroaki
H
Taniguchi
Hatsumi
H
D000397:Air Pollution; D004269:DNA, Bacterial / Q000032:analysis*; D007220:Industrial Waste / Q000032:analysis*; D009113:Muramidase / Q000494:pharmacology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D012988:Soil Microbiology; D012989:Soil Pollutants / Q000032:analysis*
D004269:DNA, Bacterial; D007220:Industrial Waste; D012989:Soil Pollutants; D012967:Sodium Dodecyl Sulfate; D009113:Muramidase