title: Illumina HiSeq 2500 sequencing; mRNA-seq for HCT116 Parental and ARID1A knockdown cells, mRNA-seq for DLD1 Parental, ARID1A knockdown and ARID1A knockout cells, mRNA-seq for COLO320DM Parental and ARID1A knockdown and ARID1A knockout cells
design: mRNA-seq for HCT116 Parental and ARID1A knockdown cells, mRNA-seq for DLD1 Parental, ARID1A knockdown and ARID1A knockout cells, mRNA-seq for COLO320DM Parental and ARID1A knockdown and ARID1A knockout cells
library_construction_protocol: Colorectal cancer cell lines were cultured in medium maintained at 37°C and 5% CO2. ARID1A knockout cells were obtained by CRISPR/Cas9 mediated gene editing. ARID1A knockdown cells were obtained by siRNA mediated knockdown. Total RNA was extracted 48 h post seeding/transfection. HCT116 cells were cultured in McCoy's 5A medium (Gibco), DLD1 and COLO320DM cells were cultured in RPMI 1640 (Gibco). All media were supplemented with 10% FBS, 1% glutamine and 100 U/mL penicillin/streptomycin. Cells were maintained at 37°C and 5% CO2.   For siRNA mediated knock down, the Lipofectamine RNAiMax transfection reagent (Invitrogen) was used according to manufacturer's instructions. The cells were harvested after 48 h. CRISPR/Cas9 mediated gene editing was used to generate ARID1A knockout colorectal cancer cell lines. The guide RNAs were cloned into a pSpCas9(BB)-2A-GFP (Addgene) plasmid. 4 μg of plasmid was transfected into the cells by electroporation using a Lonza Nucleofector. 48 h after transfection the cells were sorted as 192 single cell clones by fluorescence assisted cell sorting based on GFP-positivity. The single cell clones were expanded and selected for ARID1A knockout.    Total RNA was isolated in triplicates using the QIAzol®  lysis reagent (Qiagen) according to manufacturer's instructions. Libraries for poly(A) mRNA-sequencing were prepared using the Capture and Amplification by Tailing and Switching (CATS) RNA-seq kit (Diagenode) according to manufacturer's protocol using 50 ng total RNA as the starting material.
attributes.tag: Experimental Factor: genotype
attributes.value: ARID1A KO
attributes.tag: Experimental Factor: RNA interference
attributes.value: NT5