Human ATAD2 knockout HeLa cell line
- Advanced Validation
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- WB
Lab
Western blot - Human ATAD2 knockout HeLa cell line (AB264957)
Lanes 1- 2 : Merged signal (red and green). Green - ab176319 observed at 200 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.
ab176319 was shown to react with ATAD2 in wild-type HeLa cells in western blot. The band observed in knockout cell line ab264957 (knockout cell lysate ab257359) lane below 200kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type HeLa and ATAD2 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. ab176319 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) were incubated 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-ATAD2 antibody [EPR12730] (<a href='/en-us/products/primary-antibodies/atad2-antibody-epr12730-ab176319'>ab176319</a>) at 1/1000 dilution
Lane 1:
Wild-type HeLa cell lysate at 20 µg
Lane 2:
ATAD2 CRISPR/Cas9 edited HeLa cell lysate at 20 µg
Lane 2:
Western blot - Human ATAD2 knockout HeLa cell line (ab264957)
Predicted band size: 159 kDa
Observed band size: 200 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human ATAD2 knockout HeLa cell line (AB264957)
Homozygous : 7 bp deletion in exon 1.
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Reactivity data
Product details
We will provide viable cells that proliferate on revival.
Western blot data indicates that the CRISPR gene edit may have resulted in a truncation of the protein of interest. Please see data images.
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.
What's included?
Properties and storage information
Gene name
Gene editing type
Gene editing method
Knockout validation
Zygosity
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
ATAD2 plays a critical role in transcription regulation by remodeling chromatin structure. As part of a complex it interacts with histone acetylation marks on chromatin therefore influencing gene expression. The protein is often involved in the regulation of genes relevant to cell cycle progression. Its interaction with transcriptional regulators highlights its importance in controlling proliferation and other cellular processes.
Pathways
The role of ATAD2 becomes essential in pathways like the cell cycle and the DNA damage response. Through these pathways it interacts with key proteins such as MYC a known regulator of cell growth and it influences the expression of genes involved in cell cycle control. It is important in maintaining genomic stability by ensuring proper chromatin dynamics during DNA replication and repair.
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
Product promise
Please note: All products are 'FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC OR THERAPEUTIC PROCEDURES'.
For licensing inquiries, please contact partnerships@abcam.com