Human FZR1 knockout HEK-293T cell line
- Advanced Validation
- What is this?
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(0 Publication)
- WB
Unknown
Western blot - Human FZR1 knockout HEK-293T cell line (AB267263)
Lanes 1-4 : Merged signal (red and green). Green - ab77885 observed at 55 kDa. Red - loading control ab181602 observed at 36 kDa.
ab77885 Anti-FZR1 antibody [AR38.2] was shown to specifically react with FZR1 in wild-type HEK-293T cells. Loss of signal was observed when knockout cell line ab267263 (knockout cell lysate ab263208) was used. Wild-type and FZR1 knockout samples were subjected to SDS-PAGE. ab77885 and Anti-GAPDH antibody[EPR16891] - Loading Control (ab181602) were incubated overnight at 4° at 1 in 500 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (ab216777) and Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed (ab216772) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.
All lanes:
Western blot - Anti-FZR1 antibody [AR38.2] (<a href='/en-us/products/primary-antibodies/fzr1-antibody-ar382-ab77885'>ab77885</a>) at 1/500 dilution
Lane 1:
Wild-type HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2:
FZR1 knockout HEK-293T (Human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate at 20 µg
Lane 2:
Western blot - Human FZR1 knockout HEK-293T cell line (ab267263)
Lane 3:
HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg
Lane 4:
A431 (Human epidermoid carcinoma cell line) whole cell lysate at 20 µg
Secondary
All lanes:
Western blot - Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-680rd-preadsorbed-ab216777'>ab216777</a>) at 1/10000 dilution
Predicted band size: 55 kDa
Observed band size: 55 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human FZR1 knockout HEK-293T cell line (AB267263)
Homozygous : 1 bp insertion in exon9
- Cell Culture
Lab
Cell Culture - Human FZR1 knockout HEK-293T cell line (AB267263)
Representative images FZR1 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 a EVOS M5000 microscope.
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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
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
FZR1 helps regulate the degradation of cell cycle proteins by tagging them for destruction in a process involving the ubiquitin-proteasome system. FZR1 forms a complex with APC/C pushing the progression of the cell cycle and ensuring proper chromosome segregation. This activity prevents genomic instability by maintaining the correct order of cell cycle phases.
Pathways
Researchers place FZR1 in the cell cycle pathway particularly in regulating the mitotic exit and G1/S transition. One significant pathway involves its partnership with the cyclin B1 protein where FZR1 aids in its ubiquitination and subsequent degradation. It is also related to pathways involving the retinoblastoma protein (Rb) which adds a layer of regulation over the cycle and ensures DNA replication precision.
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