TRIB3 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 35 bp deletion in exon 2.
Images
Key facts
- Cell type
- HCT116
- Species or organism
- Human
- Tissue
- Colon
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Homozygous: 35 bp deletion in exon 2
Alternative names
C20orf97, NIPK, Neuronal cell death-inducible putative kinase, SINK, SKIP 3, TRB-3, TRIB3_HUMAN, Tribbles homolog 3, Tribbles pseudokinase 3, Tribbles3, p65-interacting inhibitor of NF-kappa-B
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TRIB3 KO cell line available to order. KO validated by Western blot. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 35 bp deletion in exon 2.
Key facts
- Cell type
- HCT116
- Species or organism
- Human
- Tissue
- Colon
- Form
- Liquid
- Knockout validation
- Sanger Sequencing, Western blot
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Homozygous: 35 bp deletion in exon 2
- Antibiotic resistance
- Puromycin 1µg/mL
- Disease
- Carcinoma
- Concentration
Properties
- Gene name
- TRIB3
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing, Western blot
- Zygosity
- Homozygous
Quality control
- STR analysis
- CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
- Biosafety level
- EU: 1 US: 1
- Adherent/suspension
- Adherent
- Gender
- Male
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
- McCoY5a + 10% FBS
- Cryopreservation medium
- Cell Freezing Medium-DMSO Serum free media, contains 8.7% DMSO in MEM supplemented with methyl cellulose.
Storage
- Shipped at conditions
- Dry Ice
- Appropriate short-term storage conditions
- -196°C
- Appropriate long-term storage conditions
- -196°C
Notes
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.
Supplementary info
- This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
TRIB3 also known as TRB3 or NIPK is a pseudokinase with a molecular mass of approximately 42 kDa. This protein does not possess catalytic activity. It functions largely through interactions with other proteins. TRIB3 is expressed in various tissues with high levels in the liver kidney and brain. The regulatory functions of TRIB3 involve modulating cellular processes such as stress response and apoptosis by interacting with signaling proteins.
- Biological function summary
The modulation of metabolic pathways and stress responses involves TRIB3 acting in several mechanisms. While TRIB3 itself is not part of a complex it exerts its effects by binding to and inhibiting the activity of other proteins notably AKT1 and several transcription factors. This modulation influences glucose metabolism and cellular response to unfolded protein stress indicating TRIB3's role as a significant regulator of cellular homeostasis.
- Pathways
The influence of TRIB3 on insulin signaling and the unfolded protein response is significant. Within the insulin signaling pathway TRIB3 interacts with AKT1 a central protein that regulates glucose uptake and cell survival. This interaction inhibits AKT1's activity impacting glucose metabolism. In the unfolded protein response pathway TRIB3 impacts stress response regulation through interactions that modulate transcriptional activity further highlighting its regulatory roles.
- Associated diseases and disorders
Alterations in TRIB3 expression and activity relate to conditions such as type 2 diabetes and cancer. In type 2 diabetes increased TRIB3 levels impair insulin signaling via AKT1 inhibition contributing to insulin resistance. In cancer the dysregulation of TRIB3 can promote survival pathways circumventing apoptosis and aiding in tumor progression. The involvement of TRIB3 in these diseases highlights its potential impact in therapeutic targeting.
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3 product images
Western blot - Human TRIB3 knockout HCT116 cell line (ab273718)
Lanes 1 - 4: Merged signal (red and green). Green - Anti-TRIB3 antibody [EPR3151Y] ab75846 observed at 40 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.Anti-TRIB3 antibody [EPR3151Y] ab75846 was shown to react with TRIB3 in wild-type HCT 116 cells in western blot with loss of signal observed in TRIB3 knockout cell line ab273718 (TRIB3 knockout cell lysate Human TRIB3 knockout HCT116 cell lysate ab275249). Wild-type and TRIB3 knockout HCT 116 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween®) before incubation with Anti-TRIB3 antibody [EPR3151Y] ab75846 and Anti-GAPDH antibody [6C5] - Loading Control ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4° at a 1 in 10000 dilution and a 1 in 20000 dilution respectively. Blots were incubated with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (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-TRIB3 antibody [EPR3151Y] (Anti-TRIB3 antibody [EPR3151Y] ab75846) at 1/10000 dilution
Lane 1: Wild-type HCT116 cell lysate at 20 µg
Lane 2: TRIB3 knockout HCT116 cell lysate at 20 µg
Lane 2: Western blot - Human TRIB3 knockout HCT116 cell line (ab273718)
Lane 3: Hap1 cell lysate at 20 µg
Lane 4: HeLa cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 40 kDa
Observed band size: 40 kDa
Sanger Sequencing - Human TRIB3 knockout HCT116 cell line (ab273718)
Allele-1: 35 bp deletion in exon 2 .
Western blot - Human TRIB3 knockout HCT116 cell line (ab273718)
False colour image of Western blot: Anti-TRIB3 antibody staining at 1/1000 dilution, shown in green; Mouse anti-Alpha Tubulin [DM1A] (Anti-alpha Tubulin antibody [DM1A] - Loading Control ab7291) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-TRIB3 antibody ab137526 was shown to bind specifically to TRIB3. A band was observed at 40 kDa in wild-type HCT 116 cell lysates with no signal observed at this size in TRIB3 knockout cell line ab273718 (knockout cell lysate Human TRIB3 knockout HCT116 cell lysate ab275249). To generate this image, wild-type and TRIB3 knockout HCT 116 cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.
All lanes: Western blot - Anti-TRIB3 antibody (Anti-TRIB3 antibody ab137526) at 1/1000 dilution
Lane 1: Wild-type HCT 116 cell lysate at 20 µg
Lane 2: Western blot - Human TRIB3 knockout HCT116 cell lysate (Human TRIB3 knockout HCT116 cell lysate ab275249) at 20 µg
SecondaryAll lanes: Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution
Performed under reducing conditions.
Predicted band size: 40 kDa
Observed band size: 40 kDa
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