GSM5256157: pri-miR-9-1 variant A; Homo sapiens; miRNA-Seq
Phenol-cholorphorm total RNA extraction 5µg of total RNA was ligated with RNA 3' adaptor using T4 RNA Ligase 2 - truncated (NEB), in the presence of RNase Inhibitor (NEB). RNA 5' adaptor was ligated using T4 RNA Ligase 1 - high concentration (NEB) and 10 mM ATP. Ligated small RNAs were reverse transcribed using SuperScript ® IV Reverse Transcriptase (Thermo-Fisher). Small RNA library cDNA was amplified and indexed using Phusion® High-Fidelity DNA polymerase (NEB). Constructs were purified in a 6% (w/v) native acrylamide gel based on the expected product size and purified by ethanol precipitation.