PDK2 KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 3 and 7 bp deletion in exon 3.
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Key facts
- Cell type
HeLa
- Species or organism
Human
- Tissue
Cervix
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 3 and 7 bp deletion in exon 3
Recommended products
PDK2 KO cell line available now. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 3 and 7 bp deletion in exon 3.
Key facts
- Cell type
HeLa
- Species or organism
Human
- Tissue
Cervix
- Form
Liquid
- Knockout validation
Sanger Sequencing, Western blot
- Mutation description
Knockout achieved by using CRISPR/Cas9, 1 bp insertion in exon 3 and 7 bp deletion in exon 3
- Disease
Adenocarcinoma
- Concentration
Properties
- Gene name
PDK2
- Gene editing type
Knockout
- Gene editing method
CRISPR technology
- Knockout validation
Sanger Sequencing, Western blot
Cell culture
- Biosafety level
EU: 2 US: 2
- Viability
~ 80%
- 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 long-term storage conditions
-196°C
Notes
Although we aim to provide customers with a homozygous clone, feasibility will be dependent on the biology of the protein. Should only heterozygous edits be achieved, you will be notified of the outcome and be asked to confirm whether the cell line is acceptable. All clones will be accompanied with DNA sequencing data, and the mutation description.
Recommended control: Human wild-type HeLa cell line (Human wild-type HeLa cell line ab255448). Please note a wild-type cell line is not automatically included with a knockout cell line order, if required please add recommended wild-type cell line at no additional cost using the code WILDTYPE-TMTK1.
Cryopreservation cell medium: Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Culture medium: DMEM (High Glucose) + 10% FBS
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 2x10E4 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 2x10E4 cells/cm2 is recommended.
• Cells should be passaged when they have achieved 80-90% confluence.
• Do not allow the cell density to exceed 7x10E4 cells/cm2.
We will provide viable cells that proliferate on revival.
This product is subject to limited use licenses from The Broad Institute, ERS Genomics Limited and Sigma-Aldrich Co. LLC, and is developed with patented technology. For full details of the licenses and 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
PDK2 also known as Pyruvate Dehydrogenase Kinase 2 is a mitochondrial protein involved in the regulation of glucose metabolism. It phosphorylates and inactivates the pyruvate dehydrogenase complex (PDC) which plays an important role in controlling the conversion of pyruvate to acetyl-CoA. PDK2 has a molecular mass of approximately 46 kDa and shows expression in various tissues including the heart liver and kidneys. The expression pattern indicates its significance in tissues with high metabolic demands.
- Biological function summary
PDK2 regulates cellular energy metabolism by phosphorylating the E1 component of PDC resulting in lowered acetyl-CoA production. PDK2 often acts as part of a regulatory complex with PDC influencing metabolic flux within the mitochondria. Its activity can alter ATP production and fatty acid synthesis impacting overall cellular function and energy homeostasis. In response to different metabolic needs the availability and activity of PDK2 modulate the balance between carbohydrate utilization and lipid oxidation.
- Pathways
PDK2 has an important role in the glucose and lipid metabolic pathways by interacting with PDC. The pathways where PDK2 is active are essential for maintaining energy homeostasis and are sensitive to different physiological conditions. PDK2's regulation of pyruvate oxidation influences the tricarboxylic acid (TCA) cycle affecting key metabolic enzymes such as pyruvate dehydrogenase phosphatase 1 (PDP1) and PDK1. Its activity is often fine-tuned by factors impacting energy needs insulin signaling and nutrient availability.
- Associated diseases and disorders
Abnormal PDK2 activity associates with metabolic conditions like diabetes and cancer. In diabetes the dysregulation of PDK2 alters the metabolic processing of carbohydrates leading to increased reliance on lipid oxidation and contributing to hyperglycemia. In cancer PDK2's role in promoting aerobic glycolysis also known as the Warburg effect supports rapid cell growth and survival. The link between PDK2 and these diseases highlights its involvement alongside other proteins such as hypoxia-inducible factor 1-alpha (HIF-1α) in promoting altered energy metabolism in pathological states.
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2 product images
Sanger Sequencing - Human PDK2 knockout HeLa cell line (ab264858)
Allele-1: 7 bp deletion in exon 3.
Sanger Sequencing - Human PDK2 knockout HeLa cell line (ab264858)
Allele-2: 1 bp insertion in exon 3.
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