Human CDK4 knockout HeLa cell line
- Advanced Validation
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CDK4 KO cell line available to order. KO validated by Western blot. Free of charge wild type control available. Knockout achieved by using CRISPR/Cas9, 139 bp insertion in exon 2 and 2 bp deletion in exon 2. To order both knockout and wild-type control cells: select '2 x 1000000 Cells/vial'. To order only knockout cells: select '1000000 Cells/vial'.
View Alternative Names
CDK4 protein, CDK4_HUMAN, CMM 3, Cell division kinase 4, Cell division protein kinase 4, Crk3, Cyclin-dependent kinase 4, MGC14458, Melanoma cutaneous malignant 3, PSK-J3, p34 cdk4
- WB
Lab
Western blot - Human CDK4 knockout HeLa cell line (AB255378)
Lanes 1- 2 : Merged signal (red and green). Green - ab108357 observed at 34 kDa. Red - Anti-Vinculin antibody [VIN-54] observed at 124 kDa.
ab108357 was shown to react with CDK4 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255378 (knockout cell lysate ab263780) was used. Wild-type HeLa and CDK4 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab108357 and Anti-Vinculin antibody [VIN-54] overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and 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-Cdk4 antibody [EPR4513-32-7] (<a href='/en-us/products/primary-antibodies/cdk4-antibody-epr4513-32-7-ab108357'>ab108357</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
CDK4 knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human CDK4 knockout HeLa cell line (ab255378)
Predicted band size: 34 kDa,55 kDa
Observed band size: 34 kDa,55 kDa
false
- WB
Lab
Western blot - Human CDK4 knockout HeLa cell line (AB255378)
Lanes 1- 2 : Merged signal (red and green). Green - ab199728 observed at 34 kDa. Red - Anti-Vinculin antibody [VIN-54] observed at 124 kDa.
ab199728 was shown to react with CDK4 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab255378 (knockout cell lysate ab263780) was used. Wild-type HeLa and CDK4 knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab199728 and Anti-Vinculin antibody [VIN-54] overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and 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-Cdk4 antibody [EPR17525] (<a href='/en-us/products/primary-antibodies/cdk4-antibody-epr17525-ab199728'>ab199728</a>) at 1/2000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
CDK4 knockout HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human CDK4 knockout HeLa cell line (ab255378)
Predicted band size: 34 kDa
Observed band size: 34 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human CDK4 knockout HeLa cell line (AB255378)
Allele-2 : 139 bp insertion in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human CDK4 knockout HeLa cell line (AB255378)
Allele-1 : 2 bp deletion in exon 2.
Reactivity data
Product details
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.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Shipped at conditions
Appropriate short-term storage conditions
Appropriate long-term storage conditions
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.
Supplementary information
This supplementary information is collated from multiple sources and compiled automatically.
Biological function summary
Cdk4 plays a fundamental role in cell cycle regulation by forming a complex with D-type cyclins. Together they phosphorylate the retinoblastoma protein (Rb) resulting in the release of E2F transcription factors which promote the expression of genes necessary for DNA replication. The Cdk4/cyclin D complex regulates the cell's commitment to enter S phase and continue cell division. This regulation is key for normal cell proliferation and tissue homeostasis. Cdk4 activity is strictly controlled by INK4 family members acting as inhibitors and adding an extra regulation layer.
Pathways
Cdk4 is an integral part of important signaling pathways like the cell cycle and PI3K/AKT pathways. In the cell cycle pathway Cdk4 relays signals downstream that drive the transition from the G1 to S phase through its interaction with cyclin D and Rb. In the PI3K/AKT pathway signaling can influence cyclin D expression indirectly affecting Cdk4 activity. Proteins such as Cdk6 closely relate to Cdk4 and often compensate or partner with Cdk4 in these pathways to maintain proper cell cycle progression.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
Target data
Complementary Products
Primary Antibodies
AB199728
20ul selling size|RabMAb|Recombinant|KO Validated
![Immunocytochemistry/ Immunofluorescence - Anti-Cdk4 antibody [EPR17525] (AB199728)](https://content.abcam.com/products/images/cdk4-antibody-epr17525-ab199728--immunocytochemistry-immunofluorescence-img77408.jpg)
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Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
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