KC-4337

293T-STAT5-Luc2-EPOR-Cell-Line

44934

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Background of 293T-STAT5-Luc2-EPOR-Cell-Line

This gene encodes the erythropoietin receptor which is a member of the cytokine receptor family. Upon erythropoietin binding, this receptor activates Jak2 tyrosine kinase which activates different intracellular pathways including: Ras/MAP kinase, phosphatidylinositol 3-kinase and STAT transcription factors. The stimulated erythropoietin receptor appears to have a role in erythroid cell survival. Defects in the erythropoietin receptor may produce erythroleukemia and familial erythrocytosis. Dysregulation of this gene may affect the growth of certain tumors. Alternate splicing results in multiple transcript variants.

Specifications

Catalog Number	KC-4337
Cell Line Name	293T-STAT5-Luc2-EPOR-Cell-Line
Host Cell Line	293T-STAT5-Luc2
Description	Stable 293T-STAT5-Luc2 cell line expressing exogenous EPOR 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 +150μg/mL Hygromycin+1μg/mL Puromycin
Selection Marker	Hygromycin and Puromycin
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

293T-STAT5-Luc2-EPOR cell line was generated using lentivirus expressing EPOR sequence.

Characterization

Figure 1: 293T-STAT5-Luc2-EPOR cells were seeded into the 96-well plate, treated with EPO for 6 hours, then readout with Bright-lite™ Luciferase Assay system.

Figure 2: Characterization of 293T-STAT5-Luc2-EPOR-1B2 cell line stable clone using western blot.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS, 150μg/mL Hygromycin and 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 1:4-1:8 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

Han C, Liu Y, Wu YY, Yang B. [Update on the role and mechanism of erythropoietin receptor in acute kidney injury and repair or fibrosis]. Sheng Li Xue Bao. 2023 Feb 25;75(1):115-129. Chinese. PMID: 36859841.
Rabadan Moraes G, Pasquier F, Marzac C, Deconinck E, Damanti CC, Leroy G, El-Khoury M, El Nemer W, Kiladjian JJ, Raslova H, Najman A, Vainchenker W, Marty C, Bellanné-Chantelot C, Plo I. An inherited gain-of-function risk allele in EPOR predisposes to familial JAK2V617F myeloproliferative neoplasms. Br J Haematol. 2022 Jul;198(1):131-136. doi: 10.1111/bjh.18165. Epub 2022 Mar 30. PMID: 35355248.
He G, Zhou J, Yang X. Expression of the erythropoietin receptor in patients with proliferative diabetic retinopathy and its correlation with postoperative visual prognosis. Ann Palliat Med. 2020 Nov;9(6):4010-4016. doi: 10.21037/apm-20-1898. Epub 2020 Nov 18. PMID: 33222459.