Bacterial and Archaeal DNA Extracted from Inoculated Experiments: Implication for the Optimization of DNA Extraction from Deep-Sea Basalts

Bacterial and Archaeal DNA Extracted from Inoculated Experiments: Implication for the Optimization of DNA Extraction from Deep-Sea Basalts

Bacterial and Archaeal DNA Extracted from Inoculated Experiments: Implication for the Optimization of DNA Extraction from Deep-Sea Basalts

Abstract

Difficulties in efficient DNA extraction from deep-sea volcanic
basalt, due to high metal concentration, complex organic matter, or
sometimes the low biomass, have hampered the understanding of
the significant biosphere both at and below the sea floor. In order to
optimize the DNA extraction from basaltic rocks, sterilized basalts
with different particle sizes and chemically synthesized goethite
were inoculated with an iron oxidizer Marinobacter aquaeolei and
an extreme halophilic archaeon Halobaculum gomorrense respectively,
and extracted with several methods. Large variations in
DNA yields by different extracting methods including FastDNAR
spin for soil kit, GeneCleanR for ancient DNA kit, UltraCleanTM
and traditional phenol-chloroform methods. Among the commercially
available kits tested here, FAST spin kit and GeneCleanR for
ancient DNA kit yield 10 times more DNA than the UltraCleanTM
kit used. In combination with FAST spin kit, skim milk greatly
enhanced the archaeal DNA yields. DNA extracting efficiency was
low with the cell number lower than 1 × 106 cells, but reached as
high as 88% with a cell number of 1 × 108 cells. On these points,
different strategies should be taken into consideration for the DNA
extraction from basalts, depending on original biomass and cell
types anticipated. FAST spin kit could provide high quality bacterial
DNA for downstream PCR whilst the combination of FAST
spin kit with skim milk would greatly enhance the archaeal DNA
yields. GeneCleanR for ancient DNA kit is also recommended for
archaeal DNA extraction from deep sea basalt due to its high yield.

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