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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 (ATCC CRL-2577) cells than in RKO-E6 (ATCC CRL-2578) cells.

1. Remove and discard culture medium.
2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.53% (w/v) EDTA solution to remove all traces of serum that contains trypsin inhibitor.
3. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
   Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
4. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting.
5. Add appropriate aliquots of the cell suspension to new culture vessels.
6. Incubate cultures at 37°C

Smith ML, et al. Involvement of the p53 tumor suppressor in repair of u.v.-type DNA damage. Oncogene 10: 1053-1059, 1995. PubMed: 7700629

Smith ML, et al. Antisense GADD45 expression results in decreased DNA repair and sensitizes cells to u.v.-irradiation or cisplatin. Oncogene 13: 2255-2263, 1996. PubMed: 8950993

Bhat MK, et al. Tumor suppressor p53 is a negative regulator in thyroid hormone receptor signaling pathways. J. Biol. Chem. 272: 28989-28993, 1997. PubMed: 9360971

Kessis TD, et al. Human papillomavirus 16 E6 expression disrupts the p53-mediated cellular response to DNA damage. Proc. Natl. Acad. Sci. USA 90: 3988-3992, 1993. PubMed: 8387205