KC-4471

CHOK1-EPHA5 Cell Line

46938

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Background of CHOK1-EPHA5 Cell Line

EPHA5 (EPH Receptor A5) is a Protein Coding gene. Diseases associated with EPHA5 include Lung Cancer and Hematologic Cancer. Among its related pathways are GPCR Pathway and ERK Signaling.

Specifications

Catalog Number	KC-4471
Cell Line Name	CHOK1-EPHA5 Cell Line
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous human EPHA5 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% RPMI1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI1640 + 10% FBS + 10μg/ml Puromycin
Selection Marker	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

Cell Line Generation

CHOK1 human EPHA5 Cell Line was generated using a lentiviral vector expressing the human EPHA5 sequence.

Characterization

Figure 1: Characterization of EPHA5 overexpression in the 293T EPHA5 stable clone using FACS.

Figure 2: Characterization of EPHA5 overexpression in the 293T EPHA5 stable clone using PCR sequencing.

Cell Resuscitation

Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10μg/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% RPMI1640 + 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

Chen Z, Chen J, Ren D, Zhang J, Yang Y, Zhang H, Mao B, Ma H. EPHA5 mutations
predict survival after immunotherapy in lung adenocarcinoma. Aging (Albany NY).
2020 Dec 3;13(1):598-618. doi: 10.18632/aging.202169. Epub 2020 Dec 3. PMID:
33288738; PMCID: PMC7834994.
Liu C, He Y, Feng X, Li J, Wang J. Expression of EPHA5 in lung adenocarcinoma
is associated with lymph node metastasis and EGFR mutation. APMIS. 2022
Jun;130(6):338-345. doi: 10.1111/apm.13222. Epub 2022 Apr 4. PMID: 35332588.
Zhang J, Zhang Z, Song W, Liu J. EPHA5 mutation impairs natural killer cell-
mediated cytotoxicity against non-small lung cancer cells and promotes cancer
cell migration and invasion. Mol Cell Probes. 2020 Aug;52:101566. doi:
1016/j.mcp.2020.101566. Epub 2020 Mar 28. PMID: 32234341.
Han X, Liu T, Zhai J, Liu C, Wang W, Nie C, Wang Q, Zhu X, Zhou H, Tian W.
Association between EPHA5 methylation status in peripheral blood
leukocytes and the risk and prognosis of gastric cancer. PeerJ. 2022 Sep
21;10:e13774. doi: 10.7717/peerj.13774. PMID: 36164608; PMCID: PMC9508887.