Human E2F1 knockout HEK-293 cell line (ab269502)
Overview
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Product name
Human E2F1 knockout HEK-293 cell line
See all E2F1 lysates -
Parental Cell Line
HEK-293 -
Organism
Human -
Mutation description
Knockout achieved by CRISPR/Cas9; X = 17 bp deletion, 1 bp insertion; Frameshift = 99% -
Passage number
<20 -
Knockout validation
Next Generation Sequencing (NGS), Western Blot (WB) -
Tested applications
Suitable for: Next Generation Sequencing, WBmore details -
Biosafety level
2 -
General notes
Recommended control: Human wild-type HEK-293 cell line (ab259776). 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. Based on cell count, seed cells in an appropriate cell culture 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.
4. Incubate the culture at 37°C incubator with 5% CO2. 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.
- A partial media change 24 hours prior to subculture may be helpful to encourage growth, if required.
- Cells should be passaged when they have achieved 80-90% confluence.
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.
We will provide viable cells that proliferate on revival.
Properties
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Number of cells
1 x 106 cells/vial, 1 mL -
Adherent /Suspension
Adherent -
Tissue
Kidney -
Cell type
epithelial -
Gender
Female -
Mycoplasma free
Yes -
Storage instructions
Shipped on Dry Ice. Store in liquid nitrogen. -
Storage buffer
Constituents: 8.7% Dimethylsulfoxide, 2% Cellulose, methyl ether -
Research areas
Target
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Function
Transcription activator that binds DNA cooperatively with dp proteins through the E2 recognition site, 5'-TTTC[CG]CGC-3' found in the promoter region of a number of genes whose products are involved in cell cycle regulation or in DNA replication. The DRTF1/E2F complex functions in the control of cell-cycle progression from G1 to S phase. E2F-1 binds preferentially RB1 protein, in a cell-cycle dependent manner. It can mediate both cell proliferation and p53-dependent apoptosis. -
Sequence similarities
Belongs to the E2F/DP family. -
Post-translational
modificationsPhosphorylated by CDK2 and cyclin A-CDK2 in the S-phase.
Acetylation stimulates DNA-binding. Enhanced under stress conditions such as DNA damage and inhibited by retinoblastoma protein pRB. Regulated by KAP1/TRIM28 which recruits HDAC1 to E2F1 resulting in deacetylation. Acetylated by P/CAF/KAT2B. -
Cellular localization
Nucleus. - Information by UniProt
Associated products
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KO cell lysates
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Related Products
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab269502 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Application | Abreviews | Notes |
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Next Generation Sequencing |
Use at an assay dependent concentration.
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WB |
Use at an assay dependent concentration.
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Notes |
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Next Generation Sequencing
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. |
Images
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17 bp deletion after Ser143 (allele 1) and 1 bp insertion after Ala146 (allele 2) of the WT protein
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All lanes : Anti-E2F1 antibody [KH95] (ab4070) at 1/500 dilution
Lane 1 : Wild-type HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
Lane 2 : E2F1 knockout HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate
Lane 3 : Hep G2 (Human liver hepatocellular carcinoma cell line) whole cell lysate
Lane 4 : U-2 OS (Human bone osteosarcoma epithelial cell line) whole cell lysate
Lane 5 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Observed band size: 55 kDa why is the actual band size different from the predicted?Lanes 1 - 5: Merged signal (red and green). Green - ab4070 observed at 55 kDa. Red - loading control ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) observed at 37 kDa.
ab4070 was shown to react with E2F1 in wild-type HEK-293 cells in Western blot with loss of signal observed in E2F1 knockout cell line ab269502 (knockout cell lysate ab281354). Wild-type HEK-293 and E2F1 knockout cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3 % milk in TBS-T (0.1 % Tween®) before incubation with ab4070 and ab181602 (Rabbit Anti-GAPDH antibody [EPR16891]) overnight at 4°C at a 1 in 500 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Mouse IgG H&L (IRDye® 800CW) preabsorbed (ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (ab216777) secondary antibodies at 1 in 20000 dilution for 1 h at room temperature before imaging.
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Knockout achieved by CRISPR/Cas9; X = 17 bp deletion, 1 bp insertion; Frameshift = 99%
Protocols
Datasheets and documents
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SDS download
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Datasheet download
References (0)
ab269502 has not yet been referenced specifically in any publications.