KC-6434

293T-FAS Cell Line

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Background of 293T-FAS Cell Line

FAS (TNFRSF6) is the sixth member of the Tumor Necrosis Factor Receptor (TNFR) superfamily (NCBI gene ID:355). It is a membrane-bound trimeric protein playing a key role in apoptosis. The FAS gene, located on the long-arm of the chromosome 10 in human and on the chromosome 19 in mice, is composed of 9 exons encoding a type-I protein. Exons 1 to 5, 6 and 7 to 9 encode the extracellular domain (ECD), the transmembrane domain (TD) and the intracellular domain (ICD) respectively. A functional domain called the Death Domain (DD), entirely encoded by the exon 9, defines the subfamily of Death Receptors such as the TNF-Receptor 1 (TNFRSF1A), DR3 (TNFRSF25), FAS (TNFSFR6), and the TRAIL receptors TRAILR1 and TRAILR2 (TNFRSF10-A and -B respectively). The mature FAS protein has 319 amino acids and a predicted molecular weight of 48 kD. FAS is expressed as a trimer mainly on activated lymphocytes and virtually in all tissues. Upon interaction with its cognate ligand FASLG, FAS triggers a biochemical cascade leading to the organized cell destruction.

Specifications

Catalog Number	KC-6434
Cell Line Name	293T-FAS Cell Line
NCBI/UniProt Accession Number	NM_000043.3
Clone Number	4#
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous FAS gene
Quantity	One vial 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-like
Incubation	37 °C with 5% CO₂
Storage	Split the saturated culture at a ratio of 1:4-1:5 every 2-3 days; seed out at about 1-3 x 10⁵ cells/mL
Doubling Time	Approximately 28 hours
Mycoplasma Status	Negative

Cell Line Generation

293T-FAS cell line was generated using a lentiviral vector expressing the human FAS sequence.

Characterization

Figure 1: Characterization of FAS overexpression in the 293T-FAS stable clone using PCR sequencing.

Figure 2: Characterization of FAS overexpression in the 293T-FAS stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM+10% FBS+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 1:4-1:5 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

Magerus A, Bercher-Brayer C, Rieux-Laucat F. The genetic landscape of the FAS pathway deficiencies. Biomed J. 2021 Aug;44(4):388-399. doi: 10.1016/j.bj.2021.06.005. Epub 2021 Jun 24. PMID: 34171534; PMCID: PMC8514852.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.