Human ATR knockout (hetero) A549 cell line
- Advanced Validation
- What is this?
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ATR KO cell line available to order. KO validated by Next Generation Sequencing, Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Heterozygous: (+/-): 49 bp deletion in exon 3.
View Alternative Names
ATR_HUMAN, Ataxia telangiectasia and Rad3 related, Ataxia telangiectasia and Rad3-related protein, FCTCS, FRAP-related protein 1, FRP-1, MEC1, MEC1 mitosis entry checkpoint 1 homolog, Protein kinase ATR, Rad3 related protein, SCKL, SCKL1, Serine/threonine-protein kinase ATR
- WB
Lab
Western blot - Human ATR knockout (hetero) A549 cell line (AB276104)
Western blot : Anti-ATR antibody staining at 1/15000 dilution shown in green; Mouse anti-Alpha Tubulin [DM1A] (ab7291) loading control staining at 1/20000 dilution shown in red.
In Western blot ab10312 was shown to bind specifically to ATR. A band was observed at 260 kDa in wild-type A549 cell lysates with a reduction in signal observed at this size in ATR heterozygous knockout cell line ab276104 (knockout cell lysate ab277987). To generate this image wild-type and ATR heterozygous knockout A549 cell lysates were analysed.
Nitrocellulose membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4°. Blots were washed in TBS-T incubated with secondary antibodies for 1 h at room temperature washed again then imaged.
Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
All lanes:
Western blot - Anti-ATR antibody (<a href='/en-us/products/primary-antibodies/atr-antibody-ab10312'>ab10312</a>) at 1/15000 dilution
Lane 1:
Wild-type A549 cell lysate at 20 µg
Lane 2:
ATR knockout A549 cell lysate at 20 µg
Lane 2:
Western blot - Human ATR knockout (hetero) A549 cell line (ab276104)
Lane 3:
HeLa cell lysate at 20 µg
Predicted band size: 301 kDa
Observed band size: 260 kDa
false
- Sanger seq
Supplier Data
Sanger Sequencing - Human ATR knockout (hetero) A549 cell line (AB276104)
Heterozygous : 84.37% 49 bp deletion in exon 3
Reactivity data
Product details
Recommended control: Human wild-type A549 cell line (ab255450). 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.
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.
- Do not allow the cell density to exceed 7x104 cells/cm2.
Culture medium
F-12K + 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
ATR plays an essential role in maintaining genomic stability. It is part of a larger protein complex that includes ATRIP (ATR-interacting protein) which helps in localizing ATR to sites of DNA damage. Once activated ATR phosphorylates various substrates including CHK1 a critical checkpoint kinase involved in cell cycle arrest during DNA repair processes. The ability of ATR to coordinate with these proteins helps cells manage DNA damage effectively and prevent genomic instability.
Pathways
ATR functions centrally in the DNA damage response and repair mechanisms particularly the ATR-Chk1 pathway. This pathway interacts closely with the ATM (Ataxia Telangiectasia Mutated) pathway which also responds to DNA damage but usually to double-strand breaks. ATR primarily acts in response to replication stress and its activation leads to the arrest of the cell cycle allowing DNA repair to occur. This cooperation between ATR and ATM highlights their complementary roles in safeguarding genomic integrity under stress.
Quality control
STR analysis
CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
Biosafety level
EU: 1 US: 1
Adherent/suspension
Adherent
Gender
Male
Target data
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