KC-5410

293T-UAS-Luc2-FGFR1-KO Cell Line

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Background of 293T-UAS-Luc2-FGFR1-KO Cell Line

The Fibroblast growth factor receptor 1 (FGFR1) gene encodes a transmembrane cytokine receptor, which comprises an extracellular region of three immunoglobulin-like domains (D1, D2 and D3), a transmembrane helix and a cytoplasmic tyrosine kinase domain. Although different isoforms have different tissue expression and varied affinity to FGFs, FGFR1-IIIc, spliced through the use of exon 8B, is the predominant isoform that carries out most of the functions of the FGFR1 gene.The downstream signaling of FGFR1 is activated by the dimerization and activation of the receptor and autophosphorylation of the tyrosine kinase domains. These downstream signaling pathways include the mitogen activated protein kinases (MAPK), the phosphatidylinositide 3 kinase/AKT (PI3K/AKT) and the phospholipase C γ (PLC). FGFR1-related signaling pathways are involved in the early development of the human embryo, and thus play an important role in gastrulation, organ specification and patterning of many tissues.

Specifications

Catalog Number	KC-5410
Cell Line Name	293T-UAS-Luc2-FGFR1-KO Cell Line
Clone Number	2B1
Host Cell Line	293T-UAS-Luc2
Description	Stable 293T-UAS-Luc2 cell clone with human FGFR1 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	DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+10%FBS+150μg/mL Hygromycin B
Selection Marker	NA
Morphology	Fibroblastoid cells growing as a monolayer
Subculture	Split saturated culture 1:4-1:5 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 28 hours
Mycoplasma Status	Negative

Cell Line Generation

293T-UAS-Luc2-FGFR1-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of 293T-UAS-Luc2-FGFR1-KO cell line stable clone using PCR sequencing.

Figure 2:Characterization of 293T-UAS-Luc2-FGFR1-KO cell line stable clone using RT-PCR sequencing.

Figure 3: Characterization of 293T-UAS-Luc2-FGFR1-KO cell line stable clone using western blot.

Cell Resuscitation

Prewarm culture medium (DMEM+10%FBS+150μg/mL Hygromycin B)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 1:4-1:5 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

Wang S, Chai X, Yan Z, Zhao S, Yang Y, Li X, Niu Y, Lin G, Su Z, Wu Z, Zhang TJ, Wu N. Novel FGFR1 Variants Are Associated with Congenital Scoliosis. Genes (Basel). 2021 Jul 24;12(8):1126. doi: 10.3390/genes12081126. PMID: 34440300; PMCID: PMC8393897.