Cell cycle time of keratinocytes (immortalized with telomerase)

Value 22.5 Hours
Organism Human Homo sapiens
Reference Shi Q, King RW. Chromosome nondisjunction yields tetraploid rather than aneuploid cells in human cell lines. Nature. 2005 Oct 13 437(7061):1038-42. Supplementary online material p. 4PubMed ID16222248
Method To obtain H2B-GFP positive N/TERT-1 cells, a retroviral vector expressing an H2B-GFP fusion protein was constructed by replacing the EGFP fragment in retroviral vector pLEGFP-N1 (BD Clontech 6059-1) with the fragment H2BGFP from pBOS-H2BGFP (BD Pharmingen 559241) cut out with Not I and Sal I. To obtain cell lines that stably express H2B-GFP, N/TERT-1 cells were grown on 6-well plates (2×10^5 per well) for 48 h and transduced by refeeding for 10 h with retroviral supernatant plus 2 µg of Polybrene (Sigma) per ml. The transduced cells were subcultured the next day into 75-cm^2 flasks. Drug selection (200 µg/ml G418) was started 2 days after transduction and continued for 2 weeks.
Comments N/TERT-1 cells found to have a normal male karyotype.
Entered by Uri M
ID 105296