Cat.No. : CSC-RT2706
Host Cell: HEK293 Target Gene: CRBN
Size: 1x10^6 cells/vial, 1mL Validation: Sequencing
| Cat. No. | CSC-RT2706 |
| Description | This cell is a stable cell line with a homozygous knockout of human CRBN using CRISPR/Cas9. |
| Target Gene | CRBN |
| Host Cell | HEK293 |
| Host Cell Species | Homo sapiens (Human) |
| Size Form | 1 vial (>10^6 cell/vial) |
| Shipping | Dry ice package |
| Storage | Liquid nirtogen |
| Revival | Rapidly thaw cells in a 37°C water bath. Transfer contents into a tube containing pre-warmed media. Centrifuge cells and seed into a 25 cm2 flask containing pre-warmed media. |
| Media Type | Cells were cultured in DMEM supplemented with 10% fetal bovine serum. |
| Growth Properties | Cells are cultured as a monolayer at 37°C in a humidified atmosphere with 5% CO2. Split at 80-90% confluence, approximately 1:3-1:6. |
| Freeze Medium | Complete medium supplemented with 10% (v/v) DMSO |
| Mycoplasma | Negative |
| Format | One frozen vial containing millions of cells |
| Storage | Liquid nitrogen |
| Safety Considerations |
The following safety precautions should be observed. 1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum. 2. No eating, drinking or smoking while handling the stable line. 3. Wash hands after handling the stable line and before leaving the lab. 4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells. 5. All waste should be considered hazardous. 6. Dispose of all liquid waste after each experiment and treat with bleach. |
| Ship | Dry ice |
Cereblon (CRBN) is a novel post-translational regulator of Cav1.2α, the main conductor of cardiac contraction in healthy and pathological states. CRBN expression is higher in heart failure (HF) patients with reduced ejection fraction (HFrEF) compared with healthy subjects. Reduced CRBN levels enhance cardiac contractility in normal and doxorubicin-induced HF mice, which may serve not only as a potential biomarker for HFrEF patients but also as a potential target for new cardioprotective therapeutic strategies. In addition, the PROTAC-based CRBN degrader TD-165 increased L-type calcium channel current (I CaL) in cardiomyocytes, showing its potential as a positive inotropic agent for the treatment of heart diseases.
In vivo and in vitro studies showed that whole-body CRBN knockout mice (CRBN−/−) and cardiac-specific knockout mice (Crbnfl/fl/Myh6Cre+ ; cKO) had enhanced cardiac contractility and increased LTCC current (I CaL) compared with their respective control groups, which was mediated by a direct interaction of CRBN with Cav1.2α. Mechanistically, the Lon domain of CRBN directly interacts with the N-terminus of Cav1.2α. Increasing CRBN levels enhanced ubiquitination and proteasomal degradation of Cav1.2α and decreased I CaL.
I CaL density was significantly increased in cardiomyocytes isolated from CRBN KO mice compared with those from WT mice (Figure 1A). A similar increase in I CaL density was observed in cardiomyocytes isolated from CRBN cKO compared with that of cWT cardiomyocytes (Figure 1B). Cav1.2α-overexpressed HEK293 CRBN knockout (HEK293 KO) had a higher I CaL density compared with Cav1.2α-overexpressed HEK293 cells (HEK293 WT) (Figure 1C). Neither activation nor inactivation of I CaL was affected by CRBN KO. Researchers further compared the protein levels of Cav1.2 among WT, CRBN KO, cWT, and CRBN cKO mice. Cav1.2α was significantly increased in both KO and CRBN cKO mouse hearts (Figure 1D).
Figure 1. CRBN knockout increases Cav1.2α activity and expression. (Park N, et al., 2022)
A: DMEM supplemented with 10% fetal bovine serum.
It is not required to add the selection antibiotics when culturing the KO cells.
A: The knockout cell product is validated by PCR amplification and Sanger Sequencing to confirm the mutation at the genomic level. Please find the detailed mutation info in the datasheet.
A: Single clonal cell.
A: No. This knockout cell product is generated using the CRISPR/Cas9 system to induce small insertions or deletions (indels) resulting in frameshift mutations. Although these frameshift mutations typically disrupt the coding gene, there is a possibility that the non-functional transcript may still be transcribed. Consequently, this could potentially yield misleading results when analyzed by RT-qPCR.
A: The cell line should be stored in liquid nitrogen for long-term preservation.
A: For most cases, we often keep at least 2 clones with different frameshift mutations. Please feel free to contact us to check if there are additional available clones.
If your question is not addressed through these resources, you can fill out the online form below and we will answer your question as soon as possible.
The cells arrive in excellent condition every time, which has been critical for our long-term studies.
Utilizing the CRBN Knockout Cell Line-HEK293 has significantly broadened our research capabilities. The knockout model has helped us gain deeper insights into the molecular mechanisms of cereblon-related pathways.
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