GSM6943880: HPr1-AR cells, veh control3 smRNA; Homo sapiens; miRNA-Seq
Total RNA was isolated via TRIzol® reagent Cell lines were treated as RNA-Seq and library preparation included ligation of 5' and 3' RNA adapters to the mature miRNAs 5ʹ‐phosphate and 3ʹ‐hydroxyl groups and 11-13 PCR cycles using a universal primer and a primer containing one of 48 index sequences, which allowed pooling of libraries and multiplex sequencing.  Prior to pooling, each individual sample's amplified cDNA construct was visualized on a DNA-HS Bioanalyzer DNA chip (Agilent Technologies) for mature miRNA and other small RNA products (140-150bp).  Successful constructs were purified using a Pippen prep (Sage Inc.), using 125 – 160 bp product size settings with separation on a 3% agarose gel. The purified samples were validated for size, purity and concentration using a DNA-HS Bioanalyzer chip.