Cat.No. : CSC-RT2717
Host Cell: A549 Target Gene: CRBN
Size: 2x10^6 cells/vial Validation: Sequencing
| Cat. No. | CSC-RT2717 |
| Description | This cell line is a stable cell line with a homozygous knockout of human CRBN using CRISPR/Cas9. |
| Target Gene | CRBN |
| Gene ID | 51185 |
| Genotype | CRBN (-/-) |
| Host Cell | A549 |
| Host Cell Species | Homo sapiens (Human) |
| Cell Type | Epithelial |
| Size | 2x10^6 cells/vial |
| SequencingResult | Allele-1: 17 bp deletion in exon Allele-2: 10 bp deletion in exon |
| 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:F12 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) has been identified as a major target for immunomodulatory drugs in multiple myeloma. Here, researchers demonstrate for the first time that CRBN expression plays a functional role in lung cancer progression by regulating autophagy through Toll-like receptors (TLR)2, TLR4, and TLR7. TLR signaling has been implicated in the induction of autophagy and plays a key role in lung cancer progression and pathogenesis. These studies suggest that CRBN could serve as an effective prognostic marker for lung cancer and provide important implications for clinical and translational lung cancer biology.
In this study, cell migration and invasion were significantly enhanced in CRBN knockout (CRBNKO) A549 cells under TLR2 (HKLM), TLR4 (LPS), or TLR7 (IQM) stimulation. Significant inhibition was observed in the presence of autophagy inhibitors 3-MA or CQ. Importantly, LC3-II levels and LC3 dots, representing autophagy induction, were significantly enhanced in CRBNKO A549 treated with TLR agonists (Figure 1A-C, LC3-II levels; Figure 1D,E, LC3 puncta). TLR4 signaling induces autophagy through BECN1 ubiquitination by TRAF6. BECN1 ubiquitination was significantly increased in CRBNKO A549 cells treated with HKLM, LPS, or IQM compared with Ctrl A549 cells (Figure 1F). The production of IL-6, CCL2, CCL20, and MMP2 is required for enhanced migration and invasion of lung cancer cells after TLR activation. In addition, CRBNKO A549 cells significantly increased IL-6, CCL2, CCL20, and MMP2 in response to three TLR agonists compared with Ctrl A549 (Figure 1G-J), while significant inhibition was observed upon co-treatment with autophagy inhibitors (Figure 1G-J). Finally, the researchers found that CRBNKO A549 cells treated with TLR agonists significantly increased single cell migration rate and colony number compared with Ctrl A549 cells. In summary, CRBN is downregulated in lung cancer cells and is associated with lung cancer progression. These studies demonstrate the association between TLR stimulation and gene signatures associated with lung cancer progression and autophagy.
Figure 1. TLR2, TLR4 and TLR7 enhance autophagy and cancer‐promoting cytokines in CRBN knockout (CRBNKO) A549 lung cancer cells. (Kim M J, et al., 2022)
A: DMEM:F12 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 Human CRBN Knockout Cell Line-A549 exhibits excellent viability and growth characteristics, which are critical for our long-term studies. The consistency batch after batch ensures that we get dependable data every time.
Using the Human CRBN Knockout Cell Line-A549 has allowed us to conduct more precise functional studies on CRBN. It’s been crucial in understanding the protein’s role in various pathways, giving us clearer insights and advancing our research objectives
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