Open AccessA new DNA extraction method by controlled alkaline treatments from consolidated subsurface sediments
Author(s) -
Kouduka Mariko,
Suko Takeshi,
Morono Yuki,
Inagaki Fumio,
Ito Kazumasa,
Suzuki Yohey
Publication year - 2012
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.2011.02437.x
Subject(s) - extraction (chemistry) , dissolution , sediment , dna extraction , dna , environmental chemistry , incubation , chemistry , microbial population biology , biomineralization , chromatography , mineralogy , bacteria , biology , biochemistry , polymerase chain reaction , gene , organic chemistry , genetics , paleontology
Microbial communities that thrive in subterranean consolidated sediments are largely unknown owing to the difficulty of extracting DNA . As this difficulty is often attributed to DNA binding onto the silica‐bearing sediment matrix, we developed a DNA extraction method for consolidated sediment from the deep subsurface in which silica minerals were dissolved by being heated under alkaline conditions. NaOH concentrations (0.07 and 0.33 N ), incubation temperatures (65 and 94 °C) and incubation times (30–90 min) before neutralization were evaluated based on the copy number of extracted prokaryotic DNA . Prokaryotic DNA was detected by quantitative PCR analysis after heating the sediment sample at 94 °C in 0.33 N NaOH solution for 50–80 min. Results of 16 S rRNA gene sequence analysis of the extracted DNA were all consistent with regard to the dominant occurrence of the metallophilic bacterium, C upriavidus metallidurans , and P seudomonas spp. Mineralogical analysis revealed that the dissolution of a silica mineral (opal‐ CT ) during alkaline treatment was maximized at 94 °C in 0.33 N NaOH solution for 50 min, which may have resulted in the release of DNA into solution. Because the optimized protocol for DNA extraction is applicable to subterranean consolidated sediments from a different locality, the method developed here has the potential to expand our understanding of the microbial community structure of the deep biosphere.