GSM9155550: RNA004_50mMNAIN3_WT; synthetic construct; OTHER
We used 600ng of in vitro transcribed (IVT) RNA for library preparation following the manufacturer's protocol for the direct RNA library preparation kit SQK-RNA004 from Oxford Nanopore Technologies. The libraries were loaded onto FLO-PRO004RA PromethION RNA flow cells and sequenced on a PromethION 2 Solo device.The library preparation steps were the same as those for SQK-RNA002 except the following: 1) 1μl of SUPERase·In™ RNase Inhibitor (Invitrogen, AM2694) is added during the RT Adapter ligation reaction, 2) 6μl of RNA Ligation (RLA) instead of 6μl of RNA Adapter (RMX) is added in the second adapter ligation reaction, 3) the adapter-ligated RNA is eluted in 33μl of RNA Elution Buffer (REB), and 4) we prepared the sample for loading by adding 100μl of Sequencing Buffer (SB) and 68μl of Library Solution (LIS) to 32μl of the library sample.