Human NR2C2 (TR4) knockout HEK-293T cell line
- Advanced Validation
- What is this?
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- WB
Lab
Western blot - Human NR2C2 (TR4) knockout HEK-293T cell line (AB266228)
Lanes 1-3 : Merged signal (red and green). Green - ab109301 observed at 67 kDa. Red - loading control ab8245 observed at 36 kDa.
ab109301 Anti-TR4 antibody [EPR1773(2)] was shown to specifically react with TR4 in wild-type HEK293T cells. The band observed in CRISPR/Cas9 edited cell line ab266228 (CRISPR/Cas9 edited cell lysate ab257563) lane below 67 kDa may represent truncated forms and cleaved fragments. This has not been investigated further. Wild-type and TR4 CRISPR/Cas9 edited samples were subjected to SDS-PAGE. ab109301 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated at room temperature for 2.5 hours 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-TR4 antibody [EPR1773(2)] (<a href='/en-us/products/primary-antibodies/tr4-antibody-epr17732-ab109301'>ab109301</a>) at 1/1000 dilution
Lane 1:
Wild-type HEK293T cell lysate at 20 µg
Lane 2:
NR2C2 CRISPR/Cas9 edited HEK293T cell lysate at 20 µg
Lane 2:
Western blot - Human NR2C2 (TR4) knockout HEK-293T cell line (ab266228)
Lane 3:
HeLa 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: 65 kDa
Observed band size: 67 kDa
false
- Sanger seq
Unknown
Sanger Sequencing - Human NR2C2 (TR4) knockout HEK-293T cell line (AB266228)
Allele-2 : Insertion of the selection cassette in exon 6.
- Sanger seq
Unknown
Sanger Sequencing - Human NR2C2 (TR4) knockout HEK-293T cell line (AB266228)
Allele-1 : 16 bp deletion in exon6
- Cell Culture
Lab
Cell Culture - Human NR2C2 (TR4) knockout HEK-293T cell line (AB266228)
Representative images NR2C2 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.
Complementary Products
![Immunoprecipitation - Anti-TR4 antibody [EPR1773(2)] (AB109301)](https://content.abcam.com/products/images/tr4-antibody-epr17732-ab109301--immunoprecipitation-img16248.jpg)
Primary Antibodies
AB109526
RabMAb|Recombinant|KO Validated|20ul selling size
![Immunocytochemistry/ Immunofluorescence - Anti-TAK1 antibody [EPR5984] (AB109526)](https://content.abcam.com/products/images/tak1-antibody-epr5984-ab109526--immunocytochemistry-immunofluorescence-img121771.jpg)
- 4
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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, 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
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
TR4 significantly influences reproductive development and metabolic processes. It plays an important role in regulating genes involved in lipid and glucose metabolism. TR4 does not function alone; it often forms complexes with other proteins to exert its effects on gene expression. The formation of these complexes enhances its ability to control a wide range of biological activities.
Pathways
TR4 actively participates in the insulin signaling and androgen receptor pathways. It modulates the action of insulin by influencing genes associated with glucose metabolism impacting energy homeostasis. In the androgen receptor pathway TR4 shares regulatory roles with proteins like AR and NR2C1 ensuring proper development and function of reproductive organs. Its involvement in these pathways exemplifies its role in maintaining metabolic and reproductive health.
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'.
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