Water and rhizosphere bacterial assemblages in long-term uranium mine water treatment systems: implications for uranium risk and bioremediation
Water samples were collected in sterile 500 mL screw-cap bottles with a total volume of 13 L, transported to the laboratory at 4 C and processed within 4872 h. For molecular-genetic analysis, 1 L of each water sample was filtered on site through sterile 0.22 m Sterivex filtersand stored at 20 C until further processing.Rhizosphere samples from the Zadni Chodov wetland were collected in sterile plastic bags and stored at 4 C until processed. Plant roots were rinsed several times with sterile distilled water and subsamples were dried for chemical characterization. For microbial isolation, roots were gently washed with phosphate-buffered saline (PBS, pH 8.0) to reduce attached soil. Consequently, 3 g of chopped roots were used for DNA extraction and cultivation. To detach rhizosphere microorganisms, roots were shaken horizontally in 20 ml of PBS solution with glass beads for 2 h at room temperature (135 rpm). The resulting suspension was used as inoculum (1 mL) and for DNA extraction. Genomic DNA was extracted from Sterivex filter cartridges using the DNeasy PowerWater Kit. DNA concentrations were quantified with a Qubit 2.0 fluorometer. For bacterial community profiling, the V4 region of the 16S rRNA gene was amplified with the primer set 515F and 802R.