KC-5790

293T-IFNAR1-KO-rat-IFNAR1-Flag Cell Line

59035

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Background of 293T-IFNAR1-KO-rat-IFNAR1-Flag Cell Line

Interferon alpha/beta receptor subunit 1 (IFNAR1) is a critical component of the type I interferon (IFN-I) receptor complex, which also includes IFNAR2. It is a transmembrane protein encoded by the IFNAR1 gene in humans and serves as the ligand-binding subunit for type I interferons, including IFN-α, IFN-β, and other related subtypes. Upon binding of interferons to IFNAR1 and IFNAR2, the receptor complex activates the JAK-STAT signaling pathway, leading to the transcription of interferon-stimulated genes (ISGs) that mediate antiviral, antiproliferative, and immunomodulatory responses. IFNAR1 plays a central role in innate immunity by enabling cells to sense viral infections and initiate host defense mechanisms. Dysregulation or mutations in IFNAR1 have been associated with increased susceptibility to viral infections and may contribute to autoimmune disorders or impaired tumor surveillance.

Specifications

Catalog Number	KC-5790
Cell Line Name	293T-IFNAR1-KO-rat-IFNAR1-Flag Cell Line
Clone Number	10#
Host Cell Line	293T-IFNAR1-KO
Description	Stable 293T-IFNAR1-KO clone expressing exogenous rat-IFNAR1 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

Cell Line Generation

293T-IFNAR1-KO-rat-IFNAR1-Flag cell line was generated using a lentiviral vector expressing the rat-IFNAR1-Flag sequence.

Characterization

Figure 1: Characterization of rat-IFNAR1 overexpression in 293T-IFNAR1-KO-rat-IFNAR1-Flag stable clones using FACS.

Figure 2: Characterization of rat-IFNAR1 expression in 293T-IFNAR1-KO-rat-IFNAR1-Flag stable clones using PCR sequencing.

Cell Resuscitation

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

Cell Freezing

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

References

1. Shemesh M, Lochte S, Piehler J, Schreiber G. IFNAR1 and IFNAR2 play distinct roles in initiating type I interferon-induced JAK-STAT signaling and activating STATs. Sci Signal. 2021 Nov 23;14(710):eabe4627. doi: 10.1126/scisignal.abe4627. Epub 2021 Nov 23. PMID: 34813358.
2.Ni W, Niu Y, Cao S, Fan C, Fan J, Zhu L, Wang X. Intermittent hypoxia exacerbates anxiety in high-fat diet-induced diabetic mice by inhibiting TREM2-regulated IFNAR1 signaling. J Neuroinflammation. 2024 Jul 2;21(1):166. doi: 10.1186/s12974-024-03160-1. PMID: 38956653; PMCID: PMC11218348.