GSM5833719: HUPEC_12_G1G2_UD2 [small RNA]; Homo sapiens; ncRNA-Seq
Cells were resuspended with QIAzol (QIAGEN, Hilden, Germany) and RNA was extracted (Zymo Direct-zol RNA miniprep kit). Libraries were size selected after 3′ ligation and 5′ linker-ligation. cDNA generated with Superscript IV reverse transcriptase was amplified with Q5 Polymerase (NEB) in a low cycle PCR (12 cycles). Pippin prep was used to remove ligated linker-linker (143–185 bp). Pilot PCR on 10% of pippin-prepped cDNA was performed to determine appropriate number of additional PCR cycles. Tapestation (Agilent) was used to determine quality and concentration of the resulting libraries.