GSM7666882: 10A EV, small RNAs; Homo sapiens; miRNA-Seq
Total RNA samples were first pretreated as following to remove some RNA modifications that interfere with small RNA-seq library construction: 3'-aminoacyl (charged)deacylation to 3'-OH for 3'-adaptor ligation, 3'-cP (2',3'-cyclic phosphate) removal to 3'-OH for 3'-adaptor ligation, 5'-OH (hydroxyl group) phosphorylation to 5'-P for 5'-adaptor ligation, m1A and m3C demethylation for efficient reverse transcription. 1) 3'-adapter ligation; 2) 5'-adapter ligation; 3) cDNA synthesis; 4) PCR amplification; 5) size selection of 134∼160bp PCR amplified fragments (corresponding to 14∼40nt small RNA size range). The libraries were denatured as single-stranded DNA molecules, captured on Illumina flow cells, amplified in situ as sequencing clusters and sequenced for 50 cycles on Illumina NextSeq 500 system per the manufacturer?s instructions.