KC-3496

CHOK1-hACVR2A Cell Line

Stable CHOK1 cell line expressing exogenous human ACVR2A gene

34428

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Background of CHOK1-hACVR2A Cell Line

ACVR2A, also known as Activin RIIA, is an activin type II receptor. On ligand binding, ACVR2A can forms a receptor complex consisting of two type II and two type I transmembrane serine/threonine kinases. ACVR2A is a receptor of Activin A, Activin B and Inhibin A. Signaling by activins and BMPs is highly promiscuous, since apart from signaling through ALK4/7, the activin type II receptors (ACVR2A and 2B) can interact also with several type I BMP receptors (ALK1/2/3/6), which can also form complexes with the type II BMP receptor, BMPRII. At present, the drugs under development for this target include KER-050, Bimagrumab and LAE-102, all of which are in the clinical verification stage.

Specifications

Catalog Number	KC-3496
Cell Line Name	CHOK1-hACVR2A Cell Line
Clone Number	24#
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous ACVR2A 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 1:4-1:8 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
In Vivo Validation	NA

Cell Line Generation

CHOK1-ACVR2A cell line was generated using a lentiviral vector expressing the ACVR2A sequence.

Characterization

Figure 1: Characterization of endogenous ACVR2A in CHO-K1 using FACS.

Figure 2: Characterization of ACVR2A overexpression in CHO-K1 human ACVR2A cell line stable clone using FACS.

Figure 3: Characterization of ACVR2A overexpression in CHO-K1 human ACVR2A cell line stable clone using FACS.

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

Monsivais D, Nagashima T, Prunskaite-Hyyryläinen R, Nozawa K, Shimada K, Tang S, Hamor C, Agno JE, Chen F, Masand RP, Young SL, Creighton CJ, DeMayo FJ, Ikawa M, Lee SJ, Matzuk MM. Endometrial receptivity and implantation require uterine BMP signaling through an ACVR2A-SMAD1/SMAD5 axis. Nat Commun. 2021 Jun 7;12(1):3386. doi: 10.1038/s41467-021-23571-5. PMID: 34099644; PMCID: PMC8184938.
Williamson RD, O'Keeffe GW, Kenny LC. Activin signalling and pre-eclampsia: from genetic risk to pre-symptomatic biomarker. Cytokine. 2015 Feb;71(2):360-5. doi: 10.1016/j.cyto.2014.11.017. Epub 2014 Dec 13. PMID: 25510903.