KC-5707

293T-NPY1R Cell Line

58521

Home » 293T-NPY1R Cell Line

Background of 293T-NPY1R Cell Line

NPY1R (Neuropeptide Y Receptor Y1) is a Protein Coding gene. This gene belongs to the G-protein-coupled receptor superfamily. The encoded transmembrane protein mediates the function of neuropeptide Y (NPY), a neurotransmitter, and peptide YY (PYY), a gastrointestinal hormone. Neuropeptide Y (NPY) is highly abundant in the brain and involved in various physiological processes related to food intake and anxiety, as well as human diseases such as obesity and cancer. The encoded receptor undergoes fast agonist-induced internalization through clathrin-coated pits and is subsequently recycled back to the cell membrane. Activation of Y1 receptors may result in mobilization of intracellular calcium and inhibition of adenylate cyclase activity. Diseases associated with NPY1R include Anxiety and Body Mass Index Quantitative Trait Locus 11.

Specifications

Catalog Number	KC-5707
Cell Line Name	293T-NPY1R Cell Line
Clone Number	5#
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous human NPY1R 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-NPY1R cell line was generated using a lentiviral vector expressing the human NPY1R sequence.

Characterization

Figure 1: Characterization of human NPY1R overexpression in the 293T-NPY1R stable clone using FACS.

Figure 2: Characterization of human NPY1R in the 293T stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM + 10% FBS + 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. Liu L, Xu Q, Cheng L, Ma C, Xiao L, Xu D, Gao Y, Wang J, Song H. NPY1R is a novel peripheral blood marker predictive of metastasis and prognosis in breast cancer patients. Oncol Lett. 2015 Feb;9(2):891-896. doi: 10.3892/ol.2014.2721. Epub 2014 Nov 20. PMID: 25624911; PMCID: PMC4301529.
2. Wang L, Rao F, Zhang K, Mahata M, Rodriguez-Flores JL, Fung MM, Waalen J, Cockburn MG, Hamilton BA, Mahata SK, O'Connor DT. Neuropeptide Y(1) Receptor NPY1R discovery of naturally occurring human genetic variants governing gene expression in cella as well as pleiotropic effects on autonomic activity and blood pressure in vivo. J Am Coll Cardiol. 2009 Sep 1;54(10):944-54. doi: 10.1016/j.jacc.2009.05.035. Erratum in: J Am Coll Cardiol. 2009 Oct 6;54(15):1482. PMID: 19712806; PMCID: PMC2792636.
3. Ouedraogo M, Lecat S, Rochdi MD, Hachet-Haas M, Matthes H, Gicquiaux H, Verrier S, Gaire M, Glasser N, Mély Y, Takeda K, Bouvier M, Galzi JL, Bucher B. Distinct motifs of neuropeptide Y receptors differentially regulate trafficking and desensitization. Traffic. 2008 Mar;9(3):305-24. doi: 10.1111/j.1600-0854.2007.00691.x. Epub 2007 Dec 14. PMID: 18088318.
4. Park, C., Kim, J., Ko, SB. et al. Structural basis of neuropeptide Y signaling through Y1 receptor. Nat Commun 13, 853 (2022).