Dual Notch1 (Variant) -YFP and Inducible Delta-mCherry Reporter CHO-K1 (hN1G4esn) Cell Line
Cat. No.
T3033
Unit
1x10⁶ cells / 1.0 ml
Price
Inquiry
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Sale of this item is subjected to the completion of a Material Transfer Agreement (MTA) by the purchasing institution.
For for-profit organizations, please contact quotes@abmgood.com for pricing.
Specifications
| Description |
The Notch-Delta signaling pathway plays a vital role in determining cell fates by mediating communication between neighboring cells during development. The Delta ligand has two important roles: it trans-activates Notch in neighboring cells, and cis-inhibits Notch in its own cell. Recent studies suggest that inhibitory same-cell interaction between the Notch receptor and its Delta ligand have significant consequences in the modes of intercellular signaling. The dual reporter cell lines can monitor the transcriptional response of Notch signaling activity owing to its stable Notch receptor expression and corresponding citrine fluorescent protein (YFP) reporter. In addition, the dual reporter cell lines have doxycycline-inducible chimeric Delta (rDII1)-mCherry fusion transmembrane protein, making these cell lines useful as a quantitative platform for analysing gene regulatory circuits in living cells and for better understanding of the network and signaling dynamics in the Notch-Delta system. The hN1 cell line (Cat. No. T3032) stably expresses the human full length Notch1 gene while the hN1G4esn has an intracellular domain of hNotch1 that is replaced with a minimal variant of the transcriptional activator Gal4 to avoid activation of endogenous Notch targets (replacing amino acids 1742 to 2556 of hNotch1 with the amino acids 1-147 and 768-881 of Gal4). Both cell lines show no detectable endogenous Notch or Delta activities and can be induced with 100 ng/ml doxycycline to express Delta-mCherry. |
| Cat. No. | T3033 |
| Name | Dual Notch1 (Variant) -YFP and Inducible Delta-mCherry Reporter CHO-K1 (hN1G4esn) Cell Line |
| Unit | 1x10⁶ cells / 1.0 ml |
| Price | Inquiry |
| Category | Stable Cell Lines |
| Cell Type | Stable Cell Lines |
| Organism | Chinese Hamster (C. griseus) |
| Tissue | Ovary |
| Donor History | Female, Adult, Chinese hamster |
| Cell Morphology | Epithelial-like |
| Growth Properties | Adherent, epithelial-like |
| Seeding Density (cells/cm2) | 20,000 - 40,000 |
| Expression | Human Notch1 |
| Propagation Method |
Use of PriCoatTMT25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. Grow cells inECM-coated culture vessels unless otherwise specified in the Propagation Requirements below.
PriGrow III (TM003) + 10% Tet System Approved FBS (tetracycline negative) + Zeocin (400 µg/ml) + Blasticidin (10 µg/ml) + Geneticin (600 µg/ml) (G271) + Hygromycin (500 µg/ml) + 1% Penicillin/Streptomycin Solution (G255), 37.0°C, 5% CO₂ |
| Growth Conditions | Use of PriCoat™ T25 Flasks (G299) or Applied Cell Extracellular Matrix (G422) is required for cell adhesion to the culture vessels. PriGrow III (TM003) + 10% Tet System Approved FBS (tetracycline negative) + Zeocin (400 µg/ml) + Blasticidin (10 µg/ml) + Geneticin (600 µg/ml) (G271) + Hygromycin (500 µg/ml) + 1% Penicillin/Streptomycin Solution (G255), 37.0°C, 5% CO₂ |
| Subculture Protocol |
Volumes given below are for a T75 flask; proportionally increase or decrease the volume as required per culture vessel size. Subculture cells once the culture vessel is 80% confluent. 1. Aspirate the culture media, and add 2-3ml of pre-warmed 0.25% Trypsin-EDTA to the culture vessel. 2. Observe the cells under a microscope to confirm detachment (typically within 2-10 minutes). Cells that are difficult to detach can be put in 37°C, for several minutes to facilitate detachment. 3. Neutralize Trypsin-EDTA by adding an equal volume of the complete growth media into the culture vessel. 4. Transfer the culture suspension into a sterile centrifuge tube, and centrifuge at 125xg for 5 minutes. The actual centrifuge duration and speed may vary depending on the cell type. 5. Aspirate the supernatant, and re-suspend the pellet with pre-warmed fresh complete growth media. Add appropriate aliquots of the cell suspension to new culture vessels, as desired. 6. Incubate the cells at the recommended conditions. |
| QC |
1. qRT-PCR was used to assess the expression of Notch mRNA levels in the hN1G4esn and hN1 cells and compared to early T-cell progenitors; 2. The Notch activity induced YFP signal were assess using IgG-Deltaext fusion proteins fixed to plates at different concentrations and time-lapse recording. |
| Application | Research Use Only. |
| Storage Condition | Vapor phase of liquid nitrogen, or below -130°C. |
| BioSafety | II |
| Caution | For Research Use Only |
| Disclaimer |
1. Sale of this item is subjected to the completion of a Material Transfer Agreement (MTA) by the purchasing individual/institution for each cell line. If you have any questions regarding this, please contact us at licensing@abmgood.com. 2. All test parameters provided in the CoA are conducted using abm's standardized culture system and The stated values may vary under the end-user's culture conditions. Please verify that the product is suitable for your studies by referencing published papers or ordering RNA (0.5 μg, Cat.# C207, $450.00) or cell lysate (100 μg, Cat.# C206, $600.00) to perform preliminary experiments, or alternatively use our Gene Expression Assay Service (Cat# C138). All sales are final. 3. We recommend live cell shipments for ease of cell transfer and this option can be requested at the time of order placement. Please note that the end-user will need to evaluate the feasibility of live cell shipment by taking into account the final destination's temperature variation and its geographical location. 4. All of abm's cell biology products are for research use ONLY and NOT for therapeutic/diagnostic applications. abm is not liable for any repercussions arising from the use of its cell biology product(s) in therapeutic/diagnostic or any other non-RUO application(s). 5. abm makes no warranties or representations as to the accuracy of the information on this site. Citations from literature are provided for informational purposes only. abm does not warrant that such information has been shown to be accurate. 6. abm warrants that cell lines shall be viable upon initiation of culture for a period of thirty (30) days after shipment and that they shall meet the specifications on the applicable abm Material Product Information sheet, certificate of analysis, and/or catalog description. Such thirty (30) day period is referred to herein as the "Warranty Period". |
| Depositor | Caltech |
Documents
Supporting Protocol
STR
There are no STR for this product yet!
FAQs
| I want to make sure these cells express my gene of interest before I decide to buy the cell line. Can you provide a sample so this can be tested? | |
|
We do not perform extensive downstream characterization, or gene expression profiling of our cell lines. However, you can check the ‘References’ tab to see if a publication is available, or you can contact us to see if RNA or cell lysates are available for your cell line of interest. Please directly contact us for further information at quotes@abmgood.com.
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| Does abm's PriGrow series medium contain antibiotics? | |
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No, the medium does not contain antibiotics and needs to be supplemented by the end-user as desired.
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| Do I need to use abm's media and Applied Cell Extracellular Matrix (G422) to culture my cells? | |
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We strongly encourage using the recommended media and culture conditions described in the "Growth Conditions" section as they have been optimized for cell growth. Other supplier's media and coating conditions have not been tested.
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| What is the difference between Applied Cell Extracellular Matrix (G422) and Collagen Coating? | |
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The main component of our Applied Cell Extracellular Matrix (G422) is type I collagen specifically. For more information on abm's Applied Cell Extracellular Matrix please visit: http://www.abmgood.com/Applied-Cell-Extracellular-Matrix-G422.html.
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| Do I have to use T25 ECM-coated flasks for growing the cells? | |
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We strongly recommend thawing cells in the T25 flasks specified on the cell-specific product page to ensure optimal post-thaw recovery of the frozen cells before testing other culture vessels.
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| If I want to plate these cells to multi-well plates (e.g. 96 well plates) or dishes, how should I prepare the plates? | |
| What percentage of Trypsin-EDTA should be used to subculture cells? Can I use Trypsin-EDTA containing phenol red? | |
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The standard concentration used for subculturing is 0.25% Trypsin-EDTA, available at abm (Cat. No. TM050). Yes, Trypsin-EDTA containing phenol red can be used as it does not interfere with trypsinization.
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| How often do I need to change the media? | |
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Media should be changed every 2-3 days or as specified within the recommended growth conditions.
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| Why are these cells classified as biosafety level II? | |
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We follow the CDC-NIH recommendations that all mammalian sourced products should be handled at the Biological Safety Level 2 to minimize exposure of potentially infectious products. This information can be found in 'Biosafety in Microbiological and Biomedical Laboratories' (1999). Your institution's Safety Officer or Technical Services will be able to make the call as to whether BioSafety Level I is possible with these cells at your site, if desired.
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| How long can I store frozen vials? | |
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Cells that are properly frozen using an effective cryoprotective agent can be stored in liquid nitrogen vapor phase (or below -130°C) indefinitely without affecting viability.
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| What is the recommended storage temperature for cells? | |
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In general, if you received:
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| Can I substitute heat-inactivated FBS with FBS or vice versa? | |
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FBS and heat-inactivated FBS are different in their composition; they cannot be substituted for one another.
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| My cells are not detaching, what method do you recommend to trypsinize the cells? | |
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| How are live cells shipped? | |
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Live cells are shipped as a live culture in a T12.5 flask. Cells are seeded at an optimal confluency to avoid metabolic waste build up during transport. In addition, we thoroughly test our cell lines for functional freeze-thaw recovery. For more information on how to handle live cells, please view our detailed instructions here (https://www.abmgood.com/uploads/document/Cell_Handling_Instructions_Upon_Arrival_150623.pdf).
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| Why are cells not attaching and forming clumps after subculturing with trypsin? | |
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Cells form clumps and display attachment issues when the trypsinization agent is not effectively neutralized. To completely neutralize the trypsin agent, add an equal amount of complete growth media prior to centrifugation. Subtle changes in culture conditions, particularly in pH and the quality of serum used in the growth medium, may also affect the amount of clumping exhibited by the cells.
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| Why are my cells forming clumps after plating? | |
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It is important to ensure cell clumps are broken up through gentle re-suspension by pipetting up and down several times after pelleting cells by centrifugation. This will ensure a homogenous single cell suspension for even plating.
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| Do I need to add the selection drug to the complete growth medium? | |
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We recommend maintaining selection pressure using the drug concentration specified in the Growth Conditions section.
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| Where can I find the CoA for my product? | |
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Certificates of analysis can be found here (https://www.abmgood.com/CoA-Library.html).
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References
- LeBon, L et al. "Fringe proteins modulate Notch-ligand cis and trans interactions to specify signaling states" Elife Sep 25:3 (2014). PubMed: 25255098 .
- Sprinzak , D et al. "Cis-interactions between Notch and Delta generate mutually exclusive signalling states" Nature 465(7294):86–90 (2010). PubMed: 20418862 .
- Selimkhanov, J et al. "Recent advances in single-cell studies of gene regulation" Curr Opin Biotechnol 23(1):34-40 (2012). PubMed: 22154220.
- Wang, X et al. "Defining single molecular forces required to activate integrin and notch signaling" Science 340(6135):991-4 (2013). PubMed: 23704575.
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