KC-5768

HCT15-ABCB1-KO Cell Line

59099

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Background of HCT15-ABCB1-KO Cell Line

Adenosine triphosphate (ATP)-binding cassette subfamily B member 1 (ABCB1), also known as permeability glycoprotein (p-gp), multidrug-resistant protein 1 (MDR1), or cluster of differentiation 243 (CD243), is an ATP-dependent drug transporter that purges foreign substances from cells. The membrane-associated protein encoded by this gene is a member of the superfamily of ATP-binding cassette (ABC) transporters. ABC proteins transport various molecules across extra- and intra-cellular membranes. ABC genes are divided into seven distinct subfamilies (ABC1, MDR/TAP, MRP, ALD, OABP, GCN20, White). This protein is a member of the MDR/TAP subfamily. Members of the MDR/TAP subfamily are involved in multidrug resistance. The protein encoded by this gene is an ATP-dependent drug efflux pump for xenobiotic compounds with broad substrate specificity. It is responsible for decreased drug accumulation in multidrug-resistant cells and often mediates the development of resistance to anticancer drugs. This protein also functions as a transporter in the blood-brain barrier. Mutations in this gene are associated with colchicine resistance and Inflammatory bowel disease 13. Alternative splicing and the use of alternative promoters results in multiple transcript variants.

Specifications

Catalog Number	KC-5768
Cell Line Name	HCT15-ABCB1-KO Cell Line
Clone Number	1A1
Host Cell Line	HCT15
Description	Stable HCT15 clone with human ABCB1 gene knockout, No.1A1
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	90% RPMI1640+10% FBS
Selection Marker	NA
Morphology	Epithelial
Subculture	Split saturated culture 1:5-1:8 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

Cell Line Generation

HCT15-ABCB1-KO cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of HCT15-ABCB1-KO cell line stable clone using PCR sequencing.

Figure 2: Characterization of HCT15-ABCB1-KO cell line stable clone using RT-PCR sequencing.

Figure 3: Characterization of HCT15-ABCB1-KO cell line stable clone using FACS.

Figure 4: Characterization of HCT15-ABCB1-KO cell line stable clone using Western blot.

Figure 5: Tumor growth and body weight curves of BALB/c nude mice inoculated with HCT15-ABCB1-KO cells (n=5).

Cell Resuscitation

Prewarm culture medium (90% 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 1:5-1:8 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

Weng HJ, Tsai TF. ABCB1 in dermatology: roles in skin diseases and their treatment. J Mol Med (Berl). 2021 Nov;99(11):1527-1538. doi: 10.1007/s00109-021-02105-y. Epub 2021 Aug 9. PMID: 34370042; PMCID: PMC8350552.
https://www.ncbi.nlm.nih.gov/gene/5243