title: GSM3324678: miR124KO.d4.3 (ZNF787); Homo sapiens; RNA-Seq
library_construction_protocol: At 4 dpi, cells were detached with TrypLE Express (12604013, Thermo Fisher Scientific), spun down, and lysed with Qiazol (Qiagen). The phases were separated with Phase Lock Gel tubes (heavy, 2 ml, 733-2478, 5PRIME) and the aqueous phase was cleaned-up with the miRNeasy Kit (Qiagen). cDNA libraries of three independent WT whole-cell RNA, ∆miR-124 whole-cell RNA, WT-AGO2 RIP, and ΔmiR-124-AGO2 RIP samples for 0 and 4 dpi were prepared using NEBNext Ultra I Directional RNA Library Prep Kit for Illumina (E7420, NEB) with 1 µg input. PolyA selection, fragmentation, first strand and second strand cDNA synthesis, and purification were performed in combination with Agencourt AMPure Kit (A63880, Beckman Coulter) and repair/dA-tailing of the cDNA. Adapters were ligated to dA-tailed cDNA and size-selected using AMPure XP Beads (Beckman Coulter). RIP samples were not polyA-selected. Index primers (Illumina) were used to index library constructs, and a PCR was performed using NEBNext Q5 2X PCR Master Mix (M0543S, NEB). Libraries were purified using Agencourt AMPure Kit. Libraries were pooled and sequenced on a HiSeq 2500 (Illumina), resulting in single-end 75 bp reads.
attributes.tag: GEO Accession
attributes.value: GSM3324678