KC-5269

HuH7-CLDN6-KO Cell Line

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Background of HuH7-CLDN6-KO Cell Line

The CLDN6 gene encodes claudin-6, a member of the claudin family of tight junction proteins, which play critical roles in maintaining cell polarity and barrier function in epithelial tissues. Claudin-6 is primarily expressed during embryonic development and is often silenced in adult tissues, but its re-expression has been observed in various cancers, including gastric, ovarian, and testicular cancers, suggesting a potential role in tumorigenesis.

Specifications

Catalog Number	KC-5269
Cell Line Name	HuH7-CLDN6-KO Cell Line
Clone Number	1A4
Host Cell Line	HuH7
Description	Stable HuH7 cell line with CLDN6 gene knockout, No.1A4
Quantity	One vial 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
Selection Marker	NA
Morphology	Epithelial
Subculture	Split saturated culture 1:5-1:10 every 1-3 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 22 hours
Mycoplasma Status	Negative

Cell Line Generation

HuH7-CLDN6-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of HuH7-CLDN6-KO Cell Line stable clone using PCR sequencing.

Figure 2: Characterization of HuH7-CLDN6-KO Cell Line stable clone using RT-PCR sequencing.

Figure 3: Characterization of HuH7-CLDN6-KO Cell Line stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (DMEM + 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 1:5-1:10 every 1-3 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

https://www.ncbi.nlm.nih.gov/gene/9074
Turksen K, Troy TC. Claudin-6: a novel tight junction molecule is developmentally regulated in mouse embryonic epithelium. Dev Dyn. 2001 Oct;222(2):292-300. doi: 10.1002/dvdy.1174. PMID: 11668606.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.