KC-3791

MDA-MB-231-CRBN-KO Cell Line

34784

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Background of MDA-MB-231-CRBN-KO Cell Line

CRBN encodes a protein related to the Lon protease protein family. It binds to membranes enriched in phosphatidylinositol 4,5-bisphosphate, and shares a conserved modular region termed the epsin NH2-terminal homology (ENTH) domain which plays a crucial role in clathrin-mediated endocytosis through an unknown target. It modifies membrane curvature and facilitates the formation of clathrin-coated invaginations. When epsin function is disrupted, clathrin-mediated endocytosis is blocked.

Specifications

Catalog Number	KC-3791
Cell Line Name	MDA-MB-231-CRBN-KO Cell Line
Clone Number	1B1
Host Cell Line	MDA-MB-231
Description	Stable MDA-MB-231 clone with human CRBN gene knockout, No.1B1
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	RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10% FBS
Selection Marker	NA
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
In Vivo Validation	NA

Cell Line Generation

MDA-MB-231-CRBN-KO-1B1 cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of MDA-MB-231-CRBN-KO-1B1 cell line stable clone using PCR sequencing.

Figure 2: Characterization of MDA-MB-231-CRBN-KO-1B1 cell line stable clone using RT-PCR sequencing.

Figure 3: Characterization of MDA-MB-231-CRBN-KO-1B1 cell line stable clone using western blot.

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 1:4-1:5 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

Wang C, Zhang Y, Wu Y, Xing D. Developments of CRBN-based PROTACs as potential therapeutic agents. Eur J Med Chem. 2021 Dec 5;225:113749. doi: 10.1016/j.ejmech.2021.113749. Epub 2021 Aug 10. PMID: 34411892.
Barankiewicz J, Salomon-Perzyński A, Misiewicz-Krzemińska I, Lech-Marańda E. CRL4CRBN E3 Ligase Complex as a Therapeutic Target in Multiple Myeloma. Cancers (Basel). 2022 Sep 16;14(18):4492. doi: 10.3390/cancers14184492. PMID: 36139651; PMCID: PMC9496858.

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.