Cat.No. : CSC-RT0663
Host Cell: HEK293T Target Gene: FGFR2
Size: 1x10^6 cells/vial, 1mL Validation: Sequencing
| Cat. No. | CSC-RT0663 |
| Description | A stable cell line with a homozygous knockout of human FGFR2 using CRISPR/Cas9. |
| Target Gene | FGFR2 |
| Host Cell | HEK293T |
| Host Cell Species | Homo sapiens (Human) |
| Shipping | 10^6 cells/tube |
| Storage | Liquid nitrogen |
| 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 |
| Gene Name | FGFR2 |
| Gene ID | 2263 |
| 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 |
NF2 is a tumor suppressor gene that is frequently mutated in malignant pleural mesothelioma (MPM). Here, we found that cell growth, clonogenic activity, migration activity, and invasion activity of the NF2-knockout human mesothelial cell line MeT-5A (NF2-KO) were significantly increased compared with NF2-WT cell clones. Complementary DNA microarray analysis clearly revealed differences in the global gene expression profiles between NF2-WT and NF2-KO cell clones. Quantitative PCR analysis and Western blot analysis showed that upregulation of fibroblast growth factor receptor 2 (FGFR2) was concurrent with increased phosphorylation levels of JNK, c-Jun, and retinoblastoma (Rb) in NF2-KO cell clones. These increases were all abolished by exogenous NF2 expression in NF2-KO clones. Furthermore, disruption of FGFR2 in NF2-KO cell clones inhibited cell proliferation as well as the phosphorylation levels of JNK, c-Jun, and Rb. These findings suggest that NF2 deficiency may play a role in tumorigenesis of human mesothelial cells by mediating FGFR2 expression. FGFR2 will be a candidate molecule for the development of therapeutic and diagnostic strategies for NF2-deficient MPM.
MTT assays showed that the cell growth rate in NF2 and FGFR2 double knockout cell clones (NF2/FGFR2-DKO) was significantly reduced compared with NF2-KO clones (Figure 1A). In contrast, there was no significant change in the growth rate between FGFR2 knockout cells (FGFR2-KO) and parental cells (Figure 1B). In addition, disruption of FGFR2 in NF2-KO cells suppressed the NF2 knockout-induced migration and wound healing activities of NF2/FGFR2-DKO cells (Figures 1C,D). In addition, Western blot analysis showed that the phosphorylation levels of JNK and c-Jun were downregulated in NF2/FGFR2-DKO clones (Figure 1E). The researchers also found that the protein level of CDK2 and the phosphorylation level of Rb were reduced in NF2/FGFR2-DKO clones (Figure 1E). These results suggest that FGFR2 may play an important role in the proliferation of NF2-mutated mesothelioma cells.
Figure 1. Knockout of FGFR2 gene retards cell proliferation in the absence of NF2 gene. (Wahiduzzaman, Md, et al. 2019)
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.
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I've been using the Human FGFR2 Knockout Cell Line - HEK293T for over six months now, and I must say, the consistency and reliability of these cells have been exceptional.
The Human FGFR2 Knockout Cell Line - HEK293T has significantly streamlined our workflow. The cells arrived in perfect condition and were ready to use right out of the box.
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