GFPu-1 (ATCC® CRL-2794)

Organism: Homo sapiens, human  /  Cell Type: epithelialtransformed with adenovirus 5 DNA  / 

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Organism Homo sapiens, human
Cell Type epithelialtransformed with adenovirus 5 DNA
Product Format frozen
Morphology epithelial
Culture Properties adherent
Biosafety Level 2 Cells containing Adenovirus viral DNA sequences
Age fetus
Applications
The cell line can be used as a reporter of proteasome activity and enables the monitoring the function of the ubiquitin-proteasome system in an intact cell.
This line was derived in July, 2000 from the human embryonic kidney line, 293. 293 cells were transfected with a reporter consisting of a short degron, CL1, fused to the COOH-terminus of green fluorescent protein (GFPu).
Storage Conditions liquid nitrogen vapor phase
Derivation
This line was derived in July, 2000 from the human embryonic kidney line, 293. 293 cells were transfected with a reporter consisting of a short degron, CL1, fused to the COOH-terminus of green fluorescent protein (GFPu). The GFPu plasmid was created by ligating an oligonucleotide encoding ACKNWFSSLSHFVIHL into the GFP-C1 plasmid (Clontech) [PubMed:11375494]. Cells contain unstable green fluorescent protein (GFP). The cell line can be used as a reporter of proteasome activity and enables the monitoring the function of the ubiquitin-proteasome system in an intact cell. Ubiquitin dependent proteolysis has a role in cell division and apoptosis due to protein aggregation.
Comments
This line was derived in July, 2000 from the human embryonic kidney line, 293. 293 cells were transfected with a reporter consisting of a short degron, CL1, fused to the COOH-terminus of green fluorescent protein (GFPu). The GFPu plasmid was created by ligating an oligonucleotide encoding ACKNWFSSLSHFVIHL into the GFP-C1 plasmid (Clontech) [PubMed:11375494]. Cells contain unstable green fluorescent protein (GFP). The cell line can be used as a reporter of proteasome activity and enables the monitoring the function of the ubiquitin-proteasome system in an intact cell. Ubiquitin dependent proteolysis has a role in cell division and apoptosis due to protein aggregation.
Complete Growth Medium The base medium for this cell line is ATCC-formulated Dulbecco's Modified Eagle's Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium: fetal bovine serum to a final concentration of 10%.
Subculturing
Protocol:
  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.
    An inoculum of 5 X 10(3) to 3 X 10(4) viable cells/cm2 is recommended. Maintain cultures at a cell concentration between 5 X 10(4) and 5 X 10(5) cells/cm2. Do not exceed 8 X 10(5) cells/cm2.
  6. Incubate cultures at 37°C.
Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:8 is recommended
Medium Renewal: Two to three times weekly
Cryopreservation
Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO
Storage temperature: liquid nitrogen vapor phase
Culture Conditions
Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Temperature: 37.0°C
Growth Conditions: Do not allow cell density to exceed 90%.
Population Doubling Time 17 hrs
Name of Depositor RR Kopito
Year of Origin July, 2000
References

Bence NF, et al. Impairment of the ubiquitin-proteasome system by protein aggregation. Science 292: 1552-1555, 2001. PubMed: 11375494

Notice: Necessary PermitsPermits

These permits may be required for shipping this product:

  • Customers located in the state of Hawaii will need to contact the Hawaii Department of Agriculture to determine if an Import Permit is required. A copy of the permit or documentation that a permit is not required must be sent to ATCC in advance of shipment.
Basic Documentation
Restrictions

This product's use is governed by the Limited Use License. For information on purchasing a license to use this product for research (for-profit entities) or commercial purposes (any entity) other than those permitted in the limited use label license, contact the Licensing Department, Life Technologies Corporation, 5791 Van Allen Way, Carlsbad, California 92008. Phone (760) 603-7200 or outlicensing@lifetech.com

References

Bence NF, et al. Impairment of the ubiquitin-proteasome system by protein aggregation. Science 292: 1552-1555, 2001. PubMed: 11375494