KC-1578

HEK293-TNFRSF1A-KO-Cell-Line

22403

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Background of HEK293-TNFRSF1A-KO-Cell-Line

"TNFRSF1A encodes a member of the TNF receptor superfamily of proteins. The encoded receptor is found in membrane-bound and soluble forms that interact with membrane-bound and soluble forms, respectively, of its ligand, tumor necrosis factor alpha. Binding of membrane-bound tumor necrosis factor alpha to the membranebound receptor induces receptor trimerization and activation, which plays a role in cell survival, apoptosis, and inflammation. Proteolytic processing of the encoded receptor results in release of the soluble form of the receptor,which can interact with free tumor necrosis factor alpha to inhibit inflammation. Mutations in this gene underlie tumor necrosis factor receptor-associated periodic syndrome (TRAPS), characterized by fever, abdominal pain and other features. Mutations in this gene may also be associated with multiple sclerosis in human patients."

Specifications

Catalog Number	KC-1578
Cell Line Name	HEK293-TNFRSF1A-KO-Cell-Line
Host Cell Line	HEK293
Description	Stable HEK293 clone with human TNFRSF1A gene knockout, No.1-1B2
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+50µg/mL Hygromycin B+50µg/mL G418
Selection Marker	N/A
Morphology	Epithelial
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 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

HEK293-TNFRSF1A-KO-1-1B2 cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of HEK293-TNFRSF1A-KO cell line stable clone using PCR sequencing.

Cell Resuscitation

Prewarm culture medium (DMEM + 10% FBS + 50µg/mL Hygromycin B + 50µg/mL G418)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

Acuna K, Choudhary A, Locatelli E, et al. Impact of Tumor Necrosis Factor Receptor 1 (TNFR1) Polymorphism on Dry Eye Disease. Biomolecules. 2023;13(2):262.
2. Tapadar P, Pal A, Ghosal N, Dutta S, Pal R. Reactive oxygen species-dependent upregulation of death receptor, tumor necrosis factor receptor 1, is responsible for theophylline-mediated cytotoxicity in MDA-MB-231 breast cancer cells. Anticancer Drugs. 2022 Sep 1;33(8):731-740.