Human RAD23A (hHR23A) knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Lab
Western blot - Human RAD23A (hHR23A) knockout HEK-293T cell line (AB266343)
Lanes 1-4 : Merged signal (red and green). Green - ab108591 observed at 50 kDa. Red - loading control ab8245 observed at 36 kDa.
ab108591 Anti-hHR23A antibody [EPR4817] was shown to specifically react with hHR23A in wild-type HEK293T cells. Loss of signal was observed when knockout cell line ab266343 (knockout cell lysate ab258163) was used. Wild-type and hHR23A knockout samples were subjected to SDS-PAGE. ab108591 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 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-hHR23A antibody [EPR4817] (<a href='/en-us/products/primary-antibodies/hhr23a-antibody-epr4817-ab108591'>ab108591</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK293T cell lysate at 20 µg
Lane 2:
RAD23A knockout HEK293T cell lysate at 20 µg
Lane 2:
Western blot - Human RAD23A (hHR23A) knockout HEK-293T cell line (ab266343)
Lane 3:
MCF7 cell lysate at 20 µg
Lane 4:
Jurkat cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution
Predicted band size: 40 kDa
Observed band size: 50 kDa
false
- Cell Culture
Unknown
Cell Culture - Human RAD23A (hHR23A) knockout HEK-293T cell line (AB266343)
Representative images of RAD23A knockout HEK293T cells, low and high confluency examples (top left and right respectively) and wild-type HEK293T cells, low and high confluency (bottom left and right respectively) showing typical adherent, epithelial-like morphology. Images were captured at 10X magnification using an EVOS M5000 microscope.
- Sanger seq
Unknown
Sanger Sequencing - Human RAD23A (hHR23A) knockout HEK-293T cell line (AB266343)
Allele-2 : 1 bp deletion in exon 2.
- Sanger seq
Unknown
Sanger Sequencing - Human RAD23A (hHR23A) knockout HEK-293T cell line (AB266343)
Allele-1 : 2 bp deletion in exon2
Complementary Products
Cell Lines & Lysates
AB266323
Human PDCD6 (ALG-2) knockout HEK-293T cell line
Advanced Validation
- 0
- 0
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 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
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
HHR23A contributes significantly to the protein ubiquitination process an essential cellular mechanism for targeting proteins for degradation. It forms part of a complex with ubiquitin aiding in the recognition and delivery of ubiquitinated proteins to the proteasome for degradation. The protein's ability to bind to both ubiquitin and the proteasome enables its dual function in proteolytic pathways positioning hHR23A as an intermediary that regulates protein turnover and stability.
Pathways
HHR23A engages in the ubiquitin-proteasome pathway and the nucleotide excision repair (NER) pathway. It cooperates with other proteins like Rad4 (XPC in humans) in NER where it functions in DNA damage recognition and repair. In the ubiquitin-proteasome pathway hHR23A interacts with the proteasomal subunit Rpn10 and other ubiquitin-like proteins. These interactions facilitate effective proteolysis of damaged or unnecessary proteins emphasizing its central role in controlling intracellular protein levels.
Quality control
STR analysis
D7S820, D5S818, TH01, D16S539, TPOX, CSF1PO, D13S317
Cell culture
Biosafety level
EU: 2 US: 2
Adherent/suspension
Adherent
Gender
Female
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