PRKAA1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided.
Images
Key facts
- Cell type
- RAW 264.7
- Species or organism
- Mouse
- Tissue
- Lymphatic
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
Alternative names
5''-AMP-activated protein kinase catalytic subunit alpha-1, AAPK1_HUMAN, ACACA kinase, AMPK subunit alpha-1, Acetyl-CoA carboxylase kinase, HMGCR kinase, Hydroxymethylglutaryl-CoA reductase kinase, PRKAA 1, Tau-protein kinase PRKAA1
Recommended products
PRKAA1 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided.
Key facts
- Cell type
- RAW 264.7
- Species or organism
- Mouse
- Tissue
- Lymphatic
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Disease
- Carcinoma
- Concentration
Properties
- Gene name
- PRKAA1
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing, Western blot
Cell culture
- Biosafety level
- EU: 2 US: 2
- Adherent/suspension
- Adherent
- Gender
- Male
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 for bath 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 all remaining cells into an ultra-low attachment 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.
5. Once confluent passage into an appropriate flask at a density of 2x105 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
- Grow in ultra-low attachment flasks.
- Do not wash or use dissociation reagent. Cells are semi-adherent. Tap flask sharply to remove adhered cells.
- All seeding densities should be based on cell counts gained by established methods.
- A guide seeding density of 2x105 cells/mL is recommended.
- Cells should be passaged when they have achieved 80-90% confluence.
- Culture medium
- DMEM + 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
PRKAA1 also known as the alpha 1 catalytic subunit of AMP-activated protein kinase (AMPK) plays an important role in cellular energy homeostasis. It has a molecular mass of approximately 63 kDa. The protein is widely expressed in various tissues including the liver skeletal muscle and brain. As a serine/threonine kinase PRKAA1 reacts to changes in cellular energy status by activating pathways that restore energy balance.
- Biological function summary
PRKAA1 acts as an energy sensor and regulator in cells. As part of the AMPK heterotrimeric complex it regulates energy metabolism by phosphorylating various substrates involved in lipid glucose and protein metabolism. PRKAA1 activation results in increased glucose uptake fatty acid oxidation and mitochondrial biogenesis while inhibiting the synthesis of fatty acids cholesterol and triglycerides. This protein plays a significant role in maintaining cellular energy homeostasis and adapting to metabolic stress.
- Pathways
PRKAA1 links to both the mTOR signaling pathway and the insulin signaling pathway. In the mTOR pathway it interacts with proteins such as TSC2 and Raptor leading to the inhibition of mTORC1 activity. In the insulin signaling pathway PRKAA1 affects the phosphorylation state of proteins involved in glucose homeostasis. PRKAA1’s regulation of these pathways highlights its importance in metabolic processes and energy balance.
- Associated diseases and disorders
PRKAA1 shows association with metabolic disorders like type 2 diabetes and obesity. In type 2 diabetes dysregulation of PRKAA1-related pathways impacts insulin sensitivity and glucose uptake contributing to disease progression. Furthermore PRKAA1 relates to cardiac diseases often involving proteins such as PGC-1α which is important for mitochondrial biogenesis and energy metabolism in heart tissues. Understanding PRKAA1’s role offers insights into potential therapeutic targets for these metabolic diseases.
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3 product images
Western blot - Mouse PRKAA1 knockout RAW 264.7 cell line (ab280055)
False colour image of Western blot: Anti-AMPK alpha 1 antibody [Y365] staining at 1/1000 dilution shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution shown in red. In Western blot Anti-AMPK alpha 1 antibody [Y365] ab32047 was shown to bind specifically to AMPK alpha 1. A band was observed at 64 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in PRKAA1 knockout cell line ab280055 (knockout cell lysate Mouse PRKAA1 knockout RAW 264.7 cell lysate ab280114). To generate this image wild-type and PRKAA1 knockout RAW 264.7 cell lysates were analysed. First samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °. Blots were washed four times in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again four times then imaged.Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.
All lanes: Western blot - Anti-AMPK alpha 1 antibody [Y365] (Anti-AMPK alpha 1 antibody [Y365] ab32047) at 1/1000 dilution
Lane 1: Wild-type RAW 264.7 cell lysate at 20 µg
Lane 2: PRKAA1 knockout RAW 264.7 cell lysate at 20 µg
Lane 2: Western blot - Mouse PRKAA1 knockout RAW 264.7 cell line (ab280055)
Lane 3: Mouse Liver cell lysate at 20 µg
Lane 4: Neuro2A cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 64 kDa
Observed band size: 64 kDa
Western blot - Mouse PRKAA1 knockout RAW 264.7 cell line (ab280055)
False colour image of Western blot: Anti-AMPK alpha 1 antibody [2B7] staining at 1/500 dilution shown in green; Rabbit Anti-GAPDH antibody [EPR16891] (Anti-GAPDH antibody [EPR16891] - Loading Control ab181602) loading control staining at 1/20000 dilution shown in red. In Western blot Anti-AMPK alpha 1 antibody [2B7] ab110036 was shown to bind specifically to AMPK alpha 1. A band was observed at 64 kDa in wild-type RAW 264.7 cell lysates with no signal observed at this size in PRKAA1 knockout cell line ab280055 (knockout cell lysate Mouse PRKAA1 knockout RAW 264.7 cell lysate ab280114). To generate this image wild-type and PRKAA1 knockout RAW 264.7 cell lysates were analysed. First samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°. Blots were washed four times in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again four times then imaged.Secondary antibodies used were Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed ab216777) at 1/20000 dilution.
All lanes: Western blot - Anti-AMPK alpha 1 antibody [2B7] (Anti-AMPK alpha 1 antibody [2B7] ab110036) at 1/500 dilution
Lane 1: Wild-type RAW 264.7 cell lysate at 20 µg
Lane 2: PRKAA1 knockout RAW 264.7 cell lysate at 20 µg
Lane 2: Western blot - Mouse PRKAA1 knockout RAW 264.7 cell line (ab280055)
Lane 3: Mouse Liver cell lysate at 20 µg
Lane 4: Neuro2A cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 64 kDa
Observed band size: 64 kDa
Sanger Sequencing - Mouse PRKAA1 knockout RAW 264.7 cell line (ab280055)
Mouse Prkaa1 KO in Raw264.7 Cells with 2 bp deletion in Exon 4
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