ARAF KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 2.
Images
Key facts
- Cell type
- HEK-293T
- Species or organism
- Human
- Tissue
- Kidney
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 2
Alternative names
A Raf proto oncogene serine/threonine protein kinase, A raf 1, ARAF_HUMAN, Oncogene Araf1, Oncogene PKS2, PKS, PKS 2, Proto-oncogene A-Raf, Proto-oncogene A-Raf-1, Proto-oncogene Pks, RAFA 1, Ras binding protein DA Raf, Serine/threonine-protein kinase A-Raf, v raf murine sarcoma 3611 viral oncogene homolog, v raf murine sarcoma 3611 viral oncogene homolog 1, v raf oncogene homolog 1 (murine sarcoma 3611 virus)
Recommended products
ARAF KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 2.
Key facts
- Cell type
- HEK-293T
- Species or organism
- Human
- Tissue
- Kidney
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp deletion in exon 2
- Concentration
Properties
- Gene name
- ARAF
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing, Western blot
- Zygosity
- Homozygous
Quality control
- STR analysis
- CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
- Biosafety level
- EU: 2 US: 2
- Adherent/suspension
- Adherent
- Gender
- Female
Handling procedures
- Initial handling guidelines
Upon arrival, the vial should be stored in liquid nitrogen vapor phase and not at -80°C. Storage at -80°C may result in loss of viability.
1. Thaw the vial in 37°C water bath for approximately 1-2 minutes.
2. Transfer the cell suspension (0.8 mL) to a 15 mL/50 mL conical sterile polypropylene centrifuge tube containing 8.4 mL pre-warmed culture medium, wash vial with an additional 0.8 mL culture medium (total volume 10 mL) to collect remaining cells, and centrifuge at 201 x g (rcf) for 5 minutes at room temperature. 10 mL represents minimum recommended dilution. 20 mL represents maximum recommended dilution.
3. Resuspend the cell pellet in 5 mL pre-warmed culture medium and count using a haemocytometer or alternative cell counting method seed all remaining cells into a T25.
4. Incubate the culture at 37°C incubator with 5% CO2. Check the culture one day after revival and continue to check until 80% confluent. Media change can be given if needed.
5. Once confluent passage into an appropriate flask at a density of 2x104 cells/cm2. Seeding density is given as a guide only and should be scaled to align with individual lab schedules. Cultures should be monitored daily.- Subculture guidelines
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x104 cells/cm2 is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
- Culture medium
- DMEM (High Glucose) + 10% FBS
- Cryopreservation medium
- Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Storage
- Shipped at conditions
- Dry Ice
- Appropriate short-term storage conditions
- -196°C
- Appropriate long-term storage conditions
- -196°C
Notes
We will provide viable cells that proliferate on revival.
This product is subject to limited use licenses from The Broad Institute and ERS Genomics Limited, and is developed with patented technology. For full details of the limited use licenses and relevant patents please refer to our limited use license and patent pages.
Supplementary info
- This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
The A-Raf protein also called ARAF is a serine/threonine-protein kinase with a molecular mass of about 68 kDa. Scientists frequently refer to it with names like A-Raf or anti A-Raf when discussing its function. A-Raf is expressed in various tissues with higher levels found in the urogenital system and the central nervous system. This protein plays an important role in cellular signal transduction processes influencing cell proliferation and differentiation through phosphorylation activity.
- Biological function summary
The A-Raf protein participates in the regulation of cell growth and survival. It forms part of the MAPK/ERK signaling cascade which is essential for transmitting mitogenic signals from the cell surface to the nucleus. A-Raf acts as a modulator within this complex impacting the overall signaling output. The protein is intimately involved in the regulation of the developmental processes influencing cell fate and function.
- Pathways
The A-Raf protein is central to the MAPK/ERK pathway a critical signaling pathway involved in the regulation of cellular responses to growth signals. This pathway includes key components like the proteins B-Raf and C-Raf with A-Raf playing specific roles in the modulation and fine-tuning of this signal transmission. It helps mediate signals that control cell division differentiation and secretion ensuring appropriate cellular responses to external stimuli.
- Associated diseases and disorders
Aberrations in A-Raf functions have been linked to cancers particularly urogenital cancers due to its role in uncontrolled cell proliferation. Faulty A-Raf activity can also contribute to neurological disorders because of its expression in the central nervous system and involvement in neuronal signaling pathways. In these contexts it can interact with proteins like B-Raf which are important for maintaining normal cellular signaling and preventing tumorigenesis and neurodegenerative disorders.
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3 product images
Western blot - Human A-Raf knockout HEK-293T cell line (ab266351)
Lanes 1-4: Merged signal (red and green). Green - Anti-A-Raf antibody [EPR16208] ab200653 observed at 68 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.Anti-A-Raf antibody [EPR16208] ab200653 Anti-A-Raf antibody [EPR16208] was shown to specifically react with A-Raf in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab266351 (knockout cell lysate Human A-Raf knockout HEK-293T cell lysate ab257838) was used. Wild-type and A-Raf knockout samples were subjected to SDS-PAGE. Anti-A-Raf antibody [EPR16208] ab200653 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4° at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes: Western blot - Anti-A-Raf antibody [EPR16208] (Anti-A-Raf antibody [EPR16208] ab200653) at 1/1000 dilution
Lane 1: Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2: A-Raf knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2: Western blot - Human A-Raf knockout HEK-293T cell line (ab266351)
Lane 3: HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 4: Raji (Human Burkitts lymphoma cell line) whole cell lysate at 20 µg
SecondaryAll lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution
Predicted band size: 68 kDa
Observed band size: 68 kDa
Cell Culture - Human A-Raf knockout HEK-293T cell line (ab266351)
Representative images A-Raf knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using a EVOS M5000 microscope.
Sanger Sequencing - Human A-Raf knockout HEK-293T cell line (ab266351)
Homozygous: 1 bp deletion in exon 2
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