KC-2107

HCT116-TP53-KO-2A3-Cell-Line

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Background of HCT116-TP53-KO-2A3-Cell-Line

TP53, known as a tumor-suppressor gene plays an important role in cell cycle, apoptosis, and genome stability, the damages of TP53 gene due to inherit factors, mutagens (chemical, radiation, or virus) were found in many cancer types.

Specifications

Catalog Number	KC-2107
Cell Line Name	HCT116-TP53-KO-2A3-Cell-Line
Host Cell Line	Human HCT116 cell line
Description	HCT116 cell line stable clone with TP53 gene knockout
Quantity	Two vials of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10%FBS
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 24 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

HCT116-TP53-KO cell line was generated using the CRISPR method.

Characterization

Cell Resuscitation

Prewarm culture medium (RPMI1640 + 10%FBS) in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI-1640 + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage

References

Vogelstein B, Lane D, Levine AJ. Nature. 2000;408:307-310. PMID: 11099028
Zauberman A, Barak Y, Ragimov N, Levy N, Oren M. EMBO J. 1993;12:2799-2808. PMID: 8334996
Van Nguyen T, Puebla-Osorio N, Pang H, Dujka ME, Zhu C. J Exp Med. 2007;204:1453-1461. PMID: 17535972
Sablina AA, Budanov AV, Ilyinskaya GV, Agapova LS, Kravchenko JE, Chumakov PM. Nat Med. 2005;11:1306-1313. PMID: 16286925