RKO-E6 Cell Line

The cells contain a stably integrated human papilloma virus(HPV) E6 oncogene under control of the cytomegalovirus(CMV) promoter. The HPV E6 oncogene causes a decrease in normal p53 levels and functions.The RKO-E6 cell line lacks appreciable functional p53.Disruption of normal p53 function in human colon carcinoma RKO cells with the human papillomavirus E6 oncoprotein results in reduced repair of u.v.-induced DNA damage and also loss of induced repair following cellular u.v.-irradiation.RKO cells contain wild-type p53 but lack endogenous human thyroid receptor nuclear receptor(h-TRbeta1). The level of p53 protein is higher in RKO cells than in RKO-E6 cells.The RKO-E6 cell line can be used together with its parental cell line, RKO to investigate the effects of p53 loss on cellular parameters such as p53 mediated transcription and apoptosis.

2

Freeze medium: 60% Basal medium+30% FBS+10% DMSO Storage temperature: Liquid nitrogen vapor phase

MEM(PM150410)+10% FBS+1% P/S(PB180120)

Adherent

Atmosphere: Air, 95%; CO2, 5% Temperature: 37°C

Every 2 to 3 days

Epithelial

1:2 to 1:4

Remove and discard culture medium. Briefly rinse the cell layer with DPBS solution to remove all traces of serum that contains trypsin inhibitor. Add 1.0 to 2.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 2 to 3 minutes). Cells that are difficult to detach may be placed at 37°C to facilitate dispersal. Add 4.0 to 6.0 mL of complete growth medium and aspirate cells by gently pipetting. Add appropriate aliquots of the cell suspension to new culture vessels.

Colon