Isolation of polymerase chain reaction-ready bacterial DNA from Lake Baikal sediments by carboxyl-functionalised magnetic polymer microspheres |
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Authors: | Spanová Alena Rittich Bohuslav Styriak Igor Styriaková Iveta Horák Daniel |
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Institution: | Masaryk University, Faculty of Science, Institute of Experimental Biology, Tvrdého 14, CZ-602 00 Brno, Czech Republic. |
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Abstract: | Carboxyl group-containing magnetic nonporous poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate) (P(HEMA-co-EDMA)) microspheres were used for the isolation of polymerase chain reaction (PCR)-ready DNA from samples of Baikal sediments. DNA was isolated using the phenol extraction method or the Soil DNA Isolation Kit. The occurrence of false-negative results in PCR caused by the presence of extracellular inhibitors in DNA samples was solved using solid phase reversible DNA immobilisation. PCR-ready DNA was reversibly adsorbed to the microspheres in the presence of 8.0% (w/v) poly(ethylene glycol) (PEG 6000) and 2.0M sodium chloride concentrations. The adsorbed DNA was released from the microspheres in a low ionic strength TE buffer. The quality of isolated DNA was checked by PCR amplification. |
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