Baker s yeast rna

Solutions of CT-DNA (Sigma Aldrich) in phosphate buffer (650 μL, 1 mM Na 2 EDTA, 10 mM Na 2 HPO 4 , 20 mM NaCl, pH 7.0) gave a ratio of UV absorbances at 260 and 280 nm, A 260 /A 280 , of 1.80-1.90, indicating that the DNA was sufficiently free from protein. 27 The concentration of CT-DNA stock solution was determined from UV absorption at 260 nm. The circular dichroism spectral titration experiments were performed by keeping the CT-DNA concentration constant (2.7 mM bases) while varying the concentration of metal complexes from 0 to 39 μM. The CD spectra were measured in 1 mm path length quartz cuvettes on a JASCO J-815 circular dichroism spectrometer. Two scans were accumulated at a scan speed of 100 nm min -1 . All CD spectra were recorded at every 0.5 nm from 200 to 350 nm. Sample temperature was maintained at 35 °C using a JASCO MCB-100 mini-circulation bath. Spectra were corrected for buffer signal.
The baker's yeast RNA (Sigma Aldrich) stock solution was also prepared in phosphate buffer in DEPC (Sigma Aldrich) treated water. The A 260 /A 280 value was 2.10, indicating the RNA was pure. 28 The initial RNA concentration was 2.3 mM (bases). The titration of ruthenium complexes and the data collection were the same as indicated for the DNA experiments.

One milligram of calf thymus (CT) DNA (Sigma-Aldrich) or baker's yeast RNA (Sigma-Aldrich) were dissolved overnight at 4 °C in 1 ml Na 2 HPO 4 buffer (50 mM, pH 7). Solutions of SM in ethanol (EtOH) or CEES in EtOH/HCl (95%, v/v/0.5%, v/v) were prepared freshly (200 × of final concentration). Final dilutions were performed in pre-warmed PBS. Solvent controls were treated with 0.5% (v/v) solvent in PBS. Incubation was performed for 1 h at room temperature (RT). DNA was precipitated with 350 μl isopropanol, centrifuged at 5000 × g for 15 min at 4 °C, and washed with 250 μl cold EtOH (70%, v/v). Pellets were dried at 37 °C and resolved in 1 ml TE buffer (10 mM Tris, 1 mM EDTA, pH 7.9). RNA precipitation was performed with 1.3 ml ice-cold EtOH per 500 μl and centrifuged at 10,000 × g for 15 min at 4 °C. Pellets were washed with 250 μl cold EtOH (70%, v/v) and centrifuged at 13,000 × g for 15 min. Pellets were dried at 37 °C and resolved in 500 μl TE buffer.