GSM3324564: KO.d4.TC.4 (TimeCourse); Homo sapiens; RNA-Seq
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 were prepared using NEBNext Ultra I Directional RNA Library Prep Kit for Illumina (E7420, NEB) with 500 ng 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). 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.
GEO Accession
GSM3324564