KC-4096

293T-FGFR3Ⅲc-Cell-Line

41801

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Background of 293T-FGFR3Ⅲc-Cell-Line

FGFR3 is a protein-coding gene that encodes a fibroblast growth factor receptor involved in bone development and maintenance. It has also been implicated in a variety of cancers and has multiple isoforms and mutations.

Specifications

Catalog Number	KC-4096
Cell Line Name	293T-FGFR3Ⅲc-Cell-Line
Host Cell Line	293T
Description	Stable HEK293T clone expressing exogenous FGFR3Ⅲc gene
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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 1μg/mL Puromycin
Selection Marker	Puromycin
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 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

293T FGFR3ⅢC cell line was generated using a lentiviral vector expressing the FGFR3ⅢC sequence.

Characterization

Figure 1: Characterization of FGFR3Ⅲc overexpression in the 293T FGFR3Ⅲc stable clone using FACS.

Figure 2: Characterization of FGFR3Ⅲc in the 293T FGFR3Ⅲc stable clone using PCR sequencing.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS and 1μg/mL puromycin)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% DMEM + 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

Chen L, Fu L, Sun J, Huang Z, Fang M, Zinkle A, Liu X, Lu J, Pan Z, Wang Y, Liang G, Li X, Chen G, Mohammadi M. Structural basis for FGF hormone signalling. Nature. 2023 Jun;618(7966):862-870. doi: 10.1038/s41586-023-06155-9. Epub 2023 Jun 7. PMID: 37286607; PMCID: PMC10284700.
Keuper M, Häring HU, Staiger H. Circulating FGF21 Levels in Human Health and Metabolic Disease. Exp Clin Endocrinol Diabetes. 2020 Nov;128(11):752-770. doi: 10.1055/a-0879-2968. Epub 2019 May 20. PMID: 31108554.
Bookout AL, de Groot MH, Owen BM, Lee S, Gautron L, Lawrence HL, Ding X, Elmquist JK, Takahashi JS, Mangelsdorf DJ, Kliewer SA. FGF21 regulates metabolism and circadian behavior by acting on the nervous system. Nat Med. 2013
Sep;19(9):1147-52. doi: 10.1038/nm.3249. Epub 2013 Aug 11. PMID: 23933984; PMCID: PMC3769420.