title: NextSeq 500 sequencing; RNA-seq of human near-haploid cell line HAP1 with CRISPR-edited frameshift mutations
design: RNA-seq of human near-haploid cell line HAP1 with CRISPR-edited frameshift mutations
library_construction_protocol: Clonal HAP1 KO clones were obtained from Horizon Discovery. Cells were grown in IMDM (GIBCO, #21980-032) supplemented with 10% FCS (GIBCO, #10270) and harvested at 70-90% confluency. Total RNA was isolated from cells using the Direct-zol 96 RNA kit (Zymo research) according to the manufacturer's protocol. In brief, 2-3 * 106 cells were lysed in 1 ml Trizol. Ethanol was added to the supernatant and RNA purified using the Zymo-spin plate. RNA integrity was checked on a Fragment Analyzer (Advanced analytical) and prepared for sequencing using the NEBNext Ultra RNA Library Preparation Kit for Illumina (NEB) according to the manufacturer's instructions. DNA was amplified by 13-14 PCR cycles and 10-11 samples were pooled before sequencin
attributes.tag: Experimental Factor: phenotype
attributes.value: BRD3 homozygous knockout