Fluoromax 3 fluorometer

Etheno M13 ssDNA (3 μM) was incubated in the absence or presence of mHOP2-MND1 (0.2 μM) in the buffer containing 33 mM Hepes (pH 7.0), 1 mM ATP, 1 mM DTT, 0.5 mM magnesium acetate for 10 min at 37 °C. Then hRAD51 (1 μM) was added to the reaction mixture to initiate nucleoprotein filament formation. Fluorescence of etheno M13 ssDNA was excited at 300 nm and monitored at 408 nm using a FluoroMax-3 fluorometer (Jobin Yvon Inc).

The drug-sensitive strain E. coli C43(DE3)ΔacrAB harbouring the ClbM-pET16b plasmid was grown to OD₆₀₀ = 0.6, and ClbM expression was induced with 500 µM isopropyl β-d-thiogalactopyranoside (IPTG) for 2 h at 37 °C. Following this, 1 ml aliquots of cells were collected by centrifugation at 50,000g and resuspended in 50 mM Tris-HCl pH 7.5. Cells were incubated with 200 µM ethidium bromide for 1 h. After incubation, the cells were isolated and washed three times with buffer, using centrifugation at 20,000g between washes to isolate the cells. The cells were then resuspended in 3% trifluoroacetic acid to lyse the cells and release the fluorescent content. The cellular debris was removed by centrifugation at 20,000g for 10 min and the fluorescence was measured on a FluoroMax-3 fluorometer (Horiba). The excitation and emission wavelengths were 485 nm and 595 nm, respectively^{20 (link)}. Transport activity was scaled to the lowest ethidium accumulation inside the cells. Three independent experiments were performed. Data were analysed using GraphPad Prism 5.0.

TEM measurements were acquired on a Hitachi H-7500 transmission electron microscope at an accelerating voltage of 75 kV. The L-QD vesicles were visualized by negative staining TEM with 1% methylamine tungstate (Supplementary Information 1). Dynamic light scattering (DLS) was used to estimate the hydrodynamic diameters of lipid vesicle samples as well as L-QD samples (Supplementary Information 2). UV–visible absorption spectra were recorded using a nanodrop spectrophotometer. The fluorescence of the samples was monitored using a Horiba FluoroMax-3 fluorometer. L-QD vesicles were imaged in aqueous solution at room temperature using a Nikon Eclipse Ti microscope driven by the Elements software package (Supplementary Information 3). Equilibrium temperature-dependent FTIR spectra were recorded on a Varian 3100 FTIR spectrometer equipped with liquid nitrogen cooled mercury cadmium telluride (MCT) detector (Supplementary Information 4). The concentration of Se species in the supernatant of the L-QD samples was measured by a VG Plasma Quad III simultaneous Inductively Coupled Plasma - Mass Spectrometer (ICP-MS).