Rapid RNA Purification using SPRI Beads

Solid-phase reversible immobilisation (SPRI) on carboxylated paramagnetic beads (Sera-Mag Magnetic SpeedBeads, from GE Healthcare) were prepared for RNA binding as described (https://openwetware.org/wiki/SPRI_bead_mix). RNA purification was performed in 96-well plates with initial lysis by the addition of 25 µl of 6GTD lysis buffer (7.08 g 10 ml⁻¹ guanidine thiocyanate [6M], made up to 8.2 ml with water, 1 ml Tris HCL [pH 8.0] and 800 µl of 1M dithiothreitol), mixed by pipetting ten times and incubation at room temperature for 1 min [26 (link)]. To this, 75 µl of 100% ethyl alcohol and 20 µl of prepared SPRI beads were added, mixed by pipetting ten times and incubated at room temperature for 5 min. The RNA-bead complex was then immobilized by placing the 96-well plate on a magnetic rack and incubated again at room temperature for 5 min. The supernatant was then discarded, and the beads washed twice in 200 µl of freshly prepared 80% ethyl alcohol (v/v) with 30 s room-temperature incubation between each wash. The beads were air-dried for 2 min at room temperature before RNA was eluted by the addition of 20 µl of nuclease-free water.