Analysis of gene expression in TET1 depleted colorectal cancer Colo320DM cells

We aimed to analyze the relationship between TET1 and aberrant CpG methylation in colorectal cancer (CRC). We established three stable TET1 knockdown clones and negative control clones of Colo320DM cells, and carried out gene expression analysis with Agilent Human Gene Expression microarray kit. RNAi-induced TET1 knockdown was accomplished using a BLOCK-iT Pol II miR RNAi Expression Vector kit (Thermo Fisher Scientific). Two sets of oligonucleotides targeting TET1 were purchased from Invitrogen and ligated into a pcDNA6.2-GW/EmGFP-miR vector (Thermo Fisher Scientific). Cells were transfected with each TET1 knockdown vector or a pcDNA6.2-GW/EmGFP-miR-neg control plasmid (Thermo Fisher Scientific), after which they were selected with 1.0 mg/ml G418. GFP-positive colonies were isolated, and knockdown efficiencies were analyzed using RT-qPCR.