DDIT3 KO cell line available to order. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2.
Images
Key facts
- Cell type
- HeLa
- Species or organism
- Human
- Tissue
- Cervix
- Form
- Liquid
- Knockout validation
- Sanger Sequencing
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2
Alternative names
C/EBP Homology Protein, C/EBP zeta, C/EBP-homologous protein, C/EBP-homologous protein 10, CCAAT/enhancer binding protein homologous protein, CHOP, CHOP-10, DDIT3_HUMAN, DNA Damage Inducible Transcript 3, DNA damage-inducible transcript 3 protein, GADD 153, Growth Arrest and DNA Damage Inducible Protein 153, Growth arrest and DNA damage-inducible protein GADD153, MGC4154
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DDIT3 KO cell line available to order. Free of charge wild type control provided. Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2.
Key facts
- Cell type
- HeLa
- Species or organism
- Human
- Tissue
- Cervix
- Form
- Liquid
- Knockout validation
- Sanger Sequencing
- Mutation description
- Knockout achieved by using CRISPR/Cas9, Homozygous: 1 bp insertion in exon 2
- Antibiotic resistance
- Puromycin 1µg/mL
- Disease
- Adenocarcinoma
- Concentration
Properties
- Gene name
- DDIT3
- Gene editing type
- Knockout
- Gene editing method
- CRISPR technology
- Knockout validation
- Sanger Sequencing
- Zygosity
- Homozygous
Quality control
- STR analysis
- CSF1PO, D13S317, D7S820, D5S818, TH01, D16S539, TPOX
Cell culture
- Biosafety level
- EU: 2 US: 2
- Adherent/suspension
- Adherent
- Gender
- Female
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.
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, 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.
Supplementary info
- This supplementary information is collated from multiple sources and compiled automatically.
- Activity summary
DDIT3 also known as CHOP (C/EBP homologous protein) is a transcription factor commonly expressed in cells under stress conditions such as endoplasmic reticulum stress. It has a molecular weight of approximately 29 kDa. As part of the system that manages cell stress responses DDIT3 expression can occur in various tissues but it is particularly notable in the context of cellular stress regulation. Researchers often use DDIT3 immunohistochemistry to study its presence and expression pattern in different tissues.
- Biological function summary
DDIT3 plays a significant role in mediating cellular stress responses and promoting apoptosis when adaptive pathways fail. DDIT3 is not typically part of a multi-subunit complex but it acts in concert with other stress-related proteins to modulate gene expression. By inducing apoptosis DDIT3 helps remove severely damaged cells maintaining overall tissue health. However excessive DDIT3 activity under prolonged stress can lead to cell loss and tissue damage.
- Pathways
DDIT3 is involved in the unfolded protein response (UPR) and endoplasmic reticulum stress pathways. It interacts with proteins such as ATF4 and PERK which are part of the mechanism that governs these pathways. DDIT3 induction contributes to the UPR pathway balancing the cell's fate between repair and apoptosis. This balance is important for cell survival under stress and avoids detrimental cellular damage.
- Associated diseases and disorders
DDIT3 has a strong connection with diabetes and neurodegenerative diseases. In diabetes DDIT3 overactivation can result in pancreatic beta-cell apoptosis worsening the disease's progression. Furthermore its involvement in neurodegenerative diseases includes its association with the accumulation of misfolded proteins exacerbating cellular stress conditions. DDIT3's interaction with proteins such as BCL2 further influences disease pathways by enhancing apoptotic signals.
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3 product images
Western blot - Human DDIT3 knockout HeLa cell line (ab265760)
False colour image of Western blot: Anti-DDIT3 antibody [9C8] staining at 5 µg/ml, shown in green; Rabbit anti-alpha Tubulin antibody [EP1332Y] (Anti-alpha Tubulin antibody [EP1332Y] - Microtubule Marker ab52866) loading control staining at 1/20000 dilution, shown in red. In Western blot, Anti-CHOP antibody [9C8] ab11419 was shown to bind specifically to DDIT3. A band was observed at 25 kDa in wild-type y cell lysates with no signal observed at this size in DDIT3 knockout cell line ab265760 (knockout cell lysate Human DDIT3 knockout HeLa cell lysate ab256889). To generate this image, wild-type and DDIT3 knockout y cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution 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-Mouse IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 800CW) preadsorbed ab216772) and Goat anti-Rabbit IgG H&L (IRDye® 680RD) preabsorbed (Goat Anti-Rabbit IgG H&L (IRDye® 680RD) preadsorbed ab216777) at 1/20000 dilution.
All lanes: Western blot - Anti-CHOP antibody [9C8] (Anti-CHOP antibody [9C8] ab11419) at 5 µg/mL
Lane 1: Wild-type HeLa Vehicle Control Tunicamycin cell lysate at 20 µg
Lane 2: Wild-type HeLa Treated Tunicamycin cell lysate at 20 µg
Lane 3: DDIT3 knockout HeLa Vehicle Control Tunicamycin cell lysate at 20 µg
Lanes 3 - 4: Western blot - Human DDIT3 knockout HeLa cell line (ab265760)
Lanes 3 - 4: Western blot - Human DDIT3 knockout HeLa cell lysate (Human DDIT3 knockout HeLa cell lysate ab256889)
Lane 4: DDIT3 knockout HeLa Treated Tunicamycin cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 19 kDa
Observed band size: 25 kDa
Cell Culture - Human DDIT3 knockout HeLa cell line (ab265760)
Representative images of DDIT3 knockout HeLa cells, low and high confluency examples (top left and right respectively) and wild-type HeLa 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 XL Core microscope.
Sanger Sequencing - Human DDIT3 knockout HeLa cell line (ab265760)
Homozygous: 1 bp insertion in exon 2.
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