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Human TACR1-FLAG Stable Cell Line-HEK293T

Cat.No.: CSC-RG0405
Cell Line Description: HEK293T-HuTACR1-FLAG cell line is a hypotriploid human cell line, which has been transfected with a human tachykinin receptor 1 (TACR1) tagged in the N-terminus with FLAG to allow stably express of the human TACR1 tagged in the N-terminus with FLAG. It is an example of a cell line transfected using our proprietary CBTGS gene screening and amplification system.
Background: NK1 (or tachykinin receptor TACR1) is a selective receptor for substance P with relatively higher affinity for neurokinin A compared to neurokinin B (100- and 500-fold lower). Neurons in lamina I of the spinal dorsal horn express the NK1 receptor for substance P and mediate hyperalgesia, an enhanced sensitivity to painful stimuli. Although substance P does not mediate the signaling of acute pain or hyperalgesia, it is essential for the full development of stress-induced analgesia and for an aggressive response to territorial challenge. It is also suggested that substance P is important for orchestrating the response of the animal to major stressors such as pain, injury or invasion of territory. Modulation of substance P activity offers a radical new approach to the management of depression, anxiety and stress. The substance P receptor is highly expressed in areas of the brain that are implicated in these behaviors.
Growth Properties: Adherent
Morphology: Epithelial
Host Cell: HEK293T
Cell Line Validation: 1. Gene expression: RT-PCR experiments determined specific expression of human TACR1.2. Protein expression: TACR1 expression in this cell line has been validated by WB.3. Functional validation: Functional assays.
Sub-type: Tachykinin
Propagation: Complete growth medium: DMEM, 10% FBS, 1 μg/mL puromycinAtmosphere: air, 95%; carbon dioxide (CO2), 5% Temperature: 37.0°C
Starting Cells From Frozen Cell Stock: 1. Remove the packaging cell lines from liquid nitrogen and carry out a quick thaw. Float the cells in the 37°C water bath for 2 minutes until nearly (80%) thawed. Once cells are thawed, it is important to dilute the cells 1:10 in growth media immediately to reduce the potentially toxic effects of the DMSO preservative on the cells.2. Clean the outside of the vial with 70% ethanol before opening.3. Remove the cells from the vial and add slowly into a 15ml conical tube containing 10 ml pre-warmed media. 4. Centrifuge for 3 minutes 1000 xg to pellet cells and remove the supernatant.5. Add 14 ml of media and transfer cells to a T25 flask or a 100 mm culture dish.6. Place the cells in the 37°C incubator with 5% CO2.7. Allow incubation for 3-4 days to reach confluence. The cells will re-attach to the surface over a period of several days in culture at 37°C.
Subculturing: 1. Remove and discard culture medium.2. Briefly rinse the cell layer with 0.25% (w/v) Trypsin-0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor.3. Add 2.0 to 3.0 ml of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.4. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.5. Add appropriate aliquots of the cell suspension to new culture vessels.Incubate cultures at 37°C.Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:6 is recommended Medium Renewal: 2 to 3 times per week
Mycoplasma: Mycoplasma Status: Negative (MycoAlert Kit)
Freeze Medium: Complete growth medium 90%; DMSO, 10%
Storage: Liquid nitrogen
Preservation: 1. Detach cells from culture dish according to the Sub-Culture Procedure.2. Resuspend cells at a density of 5 x 10^6 cells/mL in freeze medium.Note: A T-75 culture flask typically yields enough cells for preparing two frozen vials.3. Aliquot 1 mL cells into cryogenic vials.4. Place vials in a freezing container and store at –80 °C overnight.5. Transfer vials to liquid nitrogen for long term storage. If properly stored, cells should remain stable for years.
Safety Considerations: The following safety precautions should be observed.1. Use pipette aids to prevent ingestion and keep aerosols down to a minimum.2. No eating, drinking or smoking while handling the stable line.3. Wash hands after handling the stable line and before leaving the lab.4. Decontaminate work surface with disinfectant or 70% ethanol before and after working with stable cells.5. All waste should be considered hazardous.6. Dispose of all liquid waste after each experiment and treat with bleach.
Ship: Dry ice
Gene Name: TACR1 tachykinin receptor 1 [ Homo sapiens ]
Official Symbol: TACR1
Synonyms: TACR1; tachykinin receptor 1; TAC1R; substance-P receptor; NK1R; NKIR; SPR; NK-1R; NK-1 receptor; tachykinin receptor 1 (substance P receptor; neurokinin-1 receptor);
Gene ID: 6869
mRNA Refseq: NM_001058
Protein Refseq: NP_001049
MIM: 162323
UniProt ID: P25103
Chromosome Location: 2p13.1-p12
Pathway: Calcium signaling pathway, organism-specific biosystem; Calcium signaling pathway, conserved biosystem; Class A/1 (Rhodopsin-like receptors), organism-specific biosystem; G alpha (q) signalling events, organism-specific biosystem; GPCR downstream signaling, organism-specific biosystem; GPCR ligand binding, organism-specific biosystem; Measles, organism-specific biosystem;
Function: G-protein coupled receptor activity; protein binding; receptor activity; signal transducer activity; substance P receptor activity; tachykinin receptor activity;

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