Isolation of polymerase chain reaction-ready bacterial DNA from Lake Baikal sediments by carboxyl-functionalised magnetic polymer microspheres.
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.
Spanová
Alena
A
Masaryk University, Faculty of Science, Institute of Experimental Biology, Tvrdého 14, CZ-602 00 Brno, Czech Republic.
Rittich
Bohuslav
B
Styriak
Igor
I
Styriaková
Iveta
I
Horák
Daniel
D
D001407:Bacillus / Q000737:chemistry; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification*; D004587:Electrophoresis, Agar Gel; D005618:Fresh Water / Q000032:analysis; D019015:Geologic Sediments / Q000032:analysis*; D008280:Magnetics; D008863:Microspheres; D016133:Polymerase Chain Reaction; D011109:Polymethacrylic Acids / Q000737:chemistry*
D004269:DNA, Bacterial; D011109:Polymethacrylic Acids; C459242:poly(2-hydroxyethyl methacrylate-co-ethylene dimethacrylate)