KC-4252

22Rv1-CHD7-KO-2A2-Cell-Line

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Background of 22Rv1-CHD7-KO-2A2-Cell-Line

The CHD7 gene, encoding the chromodomain helicase DNA-binding protein 7, is a multifunctional protein that plays a critical role in chromatin remodeling, transcriptional regulation, and signal transduction. CHD7 is a member of the CHD family of proteins, characterized by the presence of chromodomains and a SNF2-related helicase/ATPase domain. This protein is involved in a variety of developmental processes and is essential for neural crest development and sensory organ formation.Mutations in the CHD7 gene are primarily associated with CHARGE syndrome, a complex multisystem disorder characterized by a constellation of features including coloboma of the eye, heart defects, choanal atresia, retarded growth and development, genital hypoplasia, and ear anomalies. CHARGE syndrome exhibits a high degree of phenotypic variability, even among individuals with the same CHD7 mutation.

Specifications

Catalog Number	KC-4252
Cell Line Name	22Rv1-CHD7-KO-2A2-Cell-Line
Host Cell Line	22Rv1
Description	Stable 22Rv1 clone with human CHD7 gene knockout, No.2A2
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	RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10% FBS
Selection Marker	N/A
Morphology	Epithelial
Subculture	Split saturated culture 1:3-1:6 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

22Rv1-CHD7-KO-2A2 cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of 22Rv1-CHD7-KO-2A2 cell line stable clone using PCR sequencing.

Figure 2: Characterization of 22Rv1-CHD7-KO-2A2 cell line stable clone using RT-PCR sequencing.

Cell Resuscitation

Prewarm culture medium (RPMI1640 + 10% FBS)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 1:3-1:6 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI-1640 + 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

Vissers, L. J., et al. (2004). CHARGE syndrome caused by mutations in CHD7.Nature Genetics, 36(12), 1210-1212. https://doi.org/10.1038/ng1464
Mehaffey, C. A., et al. (2015). CHD7 regulates neural crest development through direct control of the BMP pathway.Nature Communications, 6, 7559. https://doi.org/10.1038/ncomms8559
Shankar, S., et al. (2017). CHD7 regulates multiple signaling pathways during inner ear hair cell development.Developmental Biology, 424(1), 33-44. https://doi.org/10.1016/j.ydbio.2017.01.018