High-throughput extraction of SARS-CoV-2 RNA from nasopharyngeal swabs using solid-phase reverse immobilization beads

Mahesh K R Kalikiri; Mohammad Rubayet Hasan; Faheem Mirza; Thabisile Xaba; Patrick Tang; Stephan Lorenz

150 µl of VTM specimens were mixed with an equal volume of buffer RLT (Qiagen) containing 6.7 µg/µl carrier RNA (Qiagen) and incubated at RT for 5 min. 300 µl of AmpureXP paramagnetic beads (Beckman Coulter ) were added to the sample and incubated for 6 min while mixing, either with a pipette or on a shaker at 1,600 rpm. The beads were captured by incubating the plate for 8-10 min. The supernatant was discarded, and the bead pellet washed twice with 300 µl of freshly prepared 80% ethanol. During the first wash, the pellet was resuspended and incubated on the magnet for 6 minutes. No mixing was performed during the second ethanol wash. The pellet was air-dried and the absence of ethanol visually confirmed. The RNA was eluted by adding 23 µl of RSB (Illumina) and transferring 20 µl of eluate to a fresh plate.