Cat.No. : CSC-RT0517
Host Cell: Hela Target Gene: HIF1A
Size: >1x10^6 cells/vial, 1mL Validation: Sequencing
| Cat. No. | CSC-RT0517 |
| Description | This cell line is a stable cell line with a homozygous knockout of human HIF1A using CRISPR/Cas9. |
| Target Gene | HIF1A |
| Gene ID | 3091 |
| Genotype | HIF1A (-/-) |
| Host Cell | Hela |
| Host Cell Species | Homo sapiens (Human) |
| Cell Type | Epithelial |
| Size | >1x10^6 cells/vial, 1mL |
| SequencingResult | Homozygous knockout: 1 bp insertion 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 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 1:4-1:6. |
| 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 |
Hypoxia-inducible factor 1 (HIF-1) is a key transcriptional mediator of the cellular response to hypoxia and is also involved in cancer progression. Regulation of its oxygen-sensitive HIF-1α subunit involves post-translational modifications that control its stability, subcellular localization, and activity. Phosphorylation of the HIF-1α C-terminal domain by ERK1/2 promotes nuclear accumulation of HIF-1α and stimulates HIF-1 activity, whereas lack of this modification triggers nuclear export of HIF-1α and its association with mitochondria. On the other hand, modification of the N-terminal domain of HIF-1α by CK1δ attenuates HIF-1 activity by impeding the formation of HIF-1α/ARNT heterodimers.
To investigate the interplay between the two antagonistic HIF-1α phosphorylations by CK1δ and ERK1/2 and their role in HIF-1α subcellular distribution and activity, the researchers constructed a number of HIF-1α mutant forms that combined phospho-deficient and phospho-mimetic mutations at two sites of the two kinases and expressed them in HIF1A knockout HeLa cell lines. In addition, they investigated the interactome of non-nuclear HIF-1α and how it might be affected by CK1δ. The study found that modification of non-nuclear HIF-1α by CK1δ leads to its release from mitochondria and its binding to microtubules through interactions between the N-terminal part of HIF-1α and tubulin. Furthermore, these results suggest that CK1δ-stimulated binding of endogenous HIF-1α to microtubules is most pronounced during mitosis and is required for the symmetric delivery of HIF-1α to daughter nuclei during cell division.
Figure 1. CK1δ phosphomimic mutations reduce mitochondrial association of non-nuclear HIF-1α. A Fluorescence microscopy images of HIF1A knockout HeLa cells expressing GFP-HIF-1α wt-SA or GFP-HIF-1α SD-SA grown under hypoxia (1% O2). B Western blot analysis of proteins recovered from soluble and microtubule-enriched fractions of HIF1A knockout HeLa cells expressing GFP-HIF-1α SD-SA under hypoxia (1% O2) using antibodies against the indicated proteins. (Arseni, Christina, et al. 2024)
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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The Human HIF1A Knockout Cell Line-HeLa's seamless integration has not only elevated the accuracy and efficiency of my experiments but has also paved the way for pioneering advancements that are reshaping our understanding of genetics. Its impact is instrumental in redefining the scientific landscape and propelling us towards new frontiers of knowledge, revolutionizing our approach to genetic research.
The Human HIF1A Knockout Cell Line-HeLa sample is outstanding. The knockout efficiency is consistent across different passages, ensuring reliable results in downstream experiments. It is an excellent choice for researchers studying the role of HIF1A in human diseases.
Human HIF1A Knockout Cell Line-HeLa's adaptability across diverse research realms fosters a spirit of collaboration, inspiring inventive methodologies and cultivating interdisciplinary partnerships that expand the horizons of genetic science.
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