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AKT1 KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided. Knockout.

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Images

Western blot - Human AKT1 knockout HCT116 cell line (AB286570), expandable thumbnail
  • Next Generation Sequencing - Human AKT1 knockout HCT116 cell line (AB286570), expandable thumbnail

Key facts

Cell type
HCT116
Species or organism
Human
Tissue
Colon
Form
Liquid
Knockout validation
Next Generation Sequencing, Western blot
Mutation description
Knockout.

Alternative names

AKT, AKT1_HUMAN, C AKT, MGC99656, PKB, PKB-ALPHA, PRKBA, Protein Kinase B Alpha, Protein kinase B, Proto-oncogene c-Akt, RAC, RAC Serine/Threonine Protein Kinase, RAC-ALPHA, RAC-PK-alpha, RAC-alpha serine/threonine-protein kinase, Rac protein kinase alpha, V-AKT murine thymoma viral oncogene homolog 1

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AKT1 KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided. Knockout.

Key facts

Cell type
HCT116
Form
Liquid
Mutation description
Knockout.
Disease
Carcinoma
Concentration
Loading...

Properties

Gene name
AKT1
Gene editing type
Knockout
Gene editing method
CRISPR technology
Knockout validation
Next Generation Sequencing, Western blot

Quality control

STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX

Cell culture

Biosafety level
EU: 1 US: 1
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 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
McCoY5a + 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

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 HCT116 cell line (ab288559). 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.

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

AKT1 also known as Protein Kinase B (PKB) is a serine/threonine kinase involved in various cellular processes. AKT1 plays an important role in mediating signals for cell survival growth and metabolism. This protein has a molecular weight of approximately 56 kDa. AKT1 is ubiquitously expressed in many tissues including the brain heart and lungs showing its importance in multiple physiological contexts. Phosphorylation of AKT1 at serine 473 denoted as p-AKT S473 is an important modification that regulates its activity.

Biological function summary

AKT1 regulates a broad spectrum of cellular functions through signaling pathways. It is an important player in the phosphatidylinositol 3-kinase (PI3K) and AKT pathway forming part of a complex involved in promoting cell survival and growth. AKT1 interacts closely with other proteins such as mTOR influencing cellular metabolism and autophagy. The phosphorylation state of AKT1 is critical for its activity with modifications like p-AKT S473 impacting its interaction with cellular substrates.

Pathways

AKT1 participates in the PI3K/AKT/mTOR signaling cascade instrumental in cell proliferation and metabolism. AKT1 integrates signals from insulin and growth factors modulating pathways that control cell growth and glucose uptake. This pathway involves proteins like mTOR and phospho-AKT which coordinate cellular responses to external stimuli. AKT1’s phosphorylation at serine 473 and the involvement of AKT1 E17K a gain-of-function mutation influence these pathways significantly.

Associated diseases and disorders

AKT1 has associations with cancer and metabolic disorders. Dysregulation of the PI3K/AKT/mTOR pathway involving AKT1 and mTOR often results in oncogenic transformation and uncontrolled cellular proliferation. In cancer such as breast cancer AKT1 mutations including AKT1 E17K are implicated altering cell signals for growth. Additionally AKT1 is connected to metabolic disorders such as Type 2 diabetes where it affects insulin signaling and glucose metabolism highlighting its interaction with metabolic proteins like mTOR and phospho-AKT.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

2 product images

  • Western blot - Human AKT1 knockout HCT116 cell line (ab286570), expandable thumbnail

    Western blot - Human AKT1 knockout HCT116 cell line (ab286570)

    Western blot: Anti-AKT1 antibody [BLR245L] (Anti-AKT1 antibody [BLR245L] ab314110) 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 magenta. In Western blot, Anti-AKT1 antibody [BLR245L] ab314110 was shown to bind specifically to AKT1. A band was observed at 55 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in AKT1 knockout cell line. To generate this image, wild-type and AKT1 knockout HCT 116 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 °C. 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 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

    All lanes: Western blot - Anti-AKT1 antibody [BLR245L] (Anti-AKT1 antibody [BLR245L] ab314110) at 1/1000 dilution

    Lane 1: Wild-type HCT 116 cell lysate at 20 µg

    Lane 2: AKT1 knockout HCT 116 cell lysate at 20 µg

    Lane 3: HeLa cell lysate at 20 µg

    Lane 4: A549 cell lysate at 20 µg

    Lane 5: Recombinant Human AKT1 protein Recombinant Human AKT1 protein ab116412 cell lysate at 0.2 µg

    Secondary

    All lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

    Performed under reducing conditions.

    Observed band size: 55 kDa

  • Next Generation Sequencing - Human AKT1 knockout HCT116 cell line (ab286570), expandable thumbnail

    Next Generation Sequencing - Human AKT1 knockout HCT116 cell line (ab286570)

    64 bp deletion after Arg67

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Product protocols

For this product, it's our understanding that no specific protocols are required. You can:

Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.

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