## 3 Materials and Methods
### DNA Sample Preparation and 16S rRNA Gene Analysis  
Samples for total microbial DNA analysis were obtained after filtration of 1.5 litres through Millipore nitrate-cellulose filters (type GS 0.22 \(\upmu\)m) using a VFD-35 vacuum filtration device [23] including a funnel filter cell, a collector, a vacuum pump, and an ejector. Precipitates on the filters were conserved with 96% ethanol and stored at \(-20\)\({}^{\circ}\)C before DNA isolation. DNA isolation from enrichment cultures was conducted using a kit (ZymoBIOMICS(tm) DNA Miniprep Kit, Zymo Research, USA) according to the manufacturer's manual.  
DNA isolation from the filter was conducted by a kit (Meta-G-Nome(tm) DNA Isolation Kit, Epicentre, USA) according to the manufacturer's manual. In preparing libraries for amplification, variable parts of a gene of the V3-V4 region of 16S rRNA were selected: for the amplification of the V3-V4 region, the degenerate primers For341 (5'-CCTACGGGNBGCASCAG-3\") and Rev806 (5\"-GGACTACHVGGGWTCTAAT-3\") were used. For the amplification of the V4 region, the degenerate primers For515 (5\"-GTGBCACGMGCCCGGTAA-3\") and Rev806 (5\"-GGACTACHVGGGWTCTAAT-3\") were used. Amplification was performed by real-time PCR on a CFX96 Touch (Bio-Rad, USA) device using qPCRmix-HS SYBR (European, Russia).  
Denaturation, primer annealing, and chain elongation for the V3-V4 region were carried out at temperatures of 96, 54, and 72 \({}^{\circ}\)C, respectively, and the steps for the V4 region were carried out at temperatures of 96, 58, and 72 \({}^{\circ}\)C, respectively. Purification of the desired product from each batch was conducted with Agencourt AMPure XP magnetic particles (Beckman Coulter, USA). Furthermore, high-throughput sequencing was performed via the MiSeq system (Illumina, USA) using a kit (MiSeq Kit v2, 500 cycles, Illumina, USA).  
The estimation of biofilm formation on rocks was carried out by culturing 1 g of rock in 10 mL of formation fluid with a mixture of 1 g/L of glucose and sodium acetate for each solution. The analysis of the respiratory metabolism of the attached microbial biofilm was performed by MTT assay on the 20 th and 40 th days [24], followed by imaging using a laser confocal microscope.  
Confocal microscopy was performed by adding fluorescent dyes to rock samples with biofilms; wheat germ agglutinin (WGA) aggregated with the fluorescent dye AlexaFluor 488 (W11261 ThermoFisher), which binds to monosaccharides of bacterial walls and extracellular polysaccharides (EPS), was used. SYTO 11, which binds to nucleic acids, served as a dye for cell staining. Samples were analysed using a Leica SP5 confocal microscope (Leica, Germany). Images were obtained using an argon laser with a wavelength of 488 nm for the detection of WGA fluorescence and a wavelength of 594 nm for the detection of SYTO 11, and Nomarski contrast mode was used for the detection of uncoloured particles (sand). The resulting images were analysed using the Imaris 7.0.0 software package (Bitplane, Switzerland) to calculate the area of polysaccharides and nucleic acids on rock samples.  
Uranium sorption experiments were conducted on rock samples before and after biofilm formation from distillate solutions and groundwater samples; the initial uranium concentration in each case was 5 mg/L. The water/rock ratio in the experiment was 1:20. The distribution coefficients were calculated after reaching sorption equilibrium in the system (3 days for a sample with biofilm and 48 h for a biofilm-free sample). The following equation was used for the calculation:  
\[K_{d}\ =\ \frac{[U]_{i}-[U]_{eq}}{[U]_{eq}}\times\frac{V}{m} \tag{1}\]  
where (U)I is the initial concentration of uranium, (U)eq is the equilibrium concentration of uranium in the aqueous phase, V is the volume of the aqueous phase in millilitres, and \(m\) is the mass of the sample in grams. The units of measurement are K\({}_{d}\) mL/g.