127TAg (ATCC® CRL-2817)

Organism: Mus musculus, mouse  /  Cell Type: fibroblast immortalized with SV40 large T antigenSV40 large T antigen transfected  / 

Organism Mus musculus, mouse
Cell Type fibroblast immortalized with SV40 large T antigenSV40 large T antigen transfected
Product Format frozen
Morphology fibroblast
Culture Properties adherent
Biosafety Level 2 cells containing SV40 viral DNA sequences
Age 14.5 days gestation embryo
Applications
DNA repair studies
Storage Conditions liquid nitrogen vapor phase
Images
Derivation
127TAg (Pms2 null [-/-]) is a mouse embryonic fibroblast (MEF) cell line derived from Pms-2 null embryos at day 14.5 of gestation. The Pms-2 deficient mice were constructed using ES-D3 [D3] embryonic stem cells [PubMed: 9096356]. The cell line was established by transfection with an expression vector for SV40 large T antigen [PubMed: 8538772]. The cells are transgenic for lambda LIZ (Lac I/cII).
Comments
127TAg (Pms2 null [-/-]) is a mouse embryonic fibroblast (MEF) cell line derived from Pms-2 null embryos at day 14.5 of gestation. The Pms-2 deficient mice were constructed using ES-D3 [D3] embryonic stem cells [PubMed: 9096356]. The cell line was established by transfection with an expression vector for SV40 large T antigen [PubMed: 8538772]. The cells are transgenic for lambda LIZ (Lac I/cII).
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 3 X 10(3) to 1 X 10(4) viable cells/cm2 is recommended.
  6. Incubate cultures at 37°C.
Interval: Maintain cultures at a cell concentration between 3 X 10(3) and 1 X 10(4) cells/cm2.
Subcultivation Ratio: A subcultivation ratio of 1:6 to 1:8 is recommended
Medium Renewal: Every two to three days
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
Population Doubling Time 17 hours
Name of Depositor RW Sobol
Year of Origin 2000
References

Sobol RW, et al. Requirement of mammalian DNA polymerase-beta in base-excision repair. Nature 379: 183-186, 1996. PubMed: 8538772

Sobol RW, et al. Base excision repair intermediates induce p53-independent cytotoxic and genotoxic responses. J. Biol. Chem. 278: 39951-39959, 2003. PubMed: 12882965

Narayanan L, et al. Elevated levels of mutation in multiple tissues of mice deficient in the DNA mismatch repair gene Pms2. Proc. Natl. Acad. Sci. USA 94: 122-127, 1997. PubMed: 9096356

Sobol RW, et al. Mutations associated with base excision repair deficiency and methylation-induced genotoxic stress. Proc. Natl. Acad. Sci. USA 99: 6860-6865, 2002. PubMed: 11983862

Sobol RW, et al. The lyase activity of the DNA repair protein beta-polymerase protects from DNA-damage-induced cytotoxicity. Nature 405: 807-810, 2000. PubMed: 10866204

Basic Documentation
Other Documentation
References

Sobol RW, et al. Requirement of mammalian DNA polymerase-beta in base-excision repair. Nature 379: 183-186, 1996. PubMed: 8538772

Sobol RW, et al. Base excision repair intermediates induce p53-independent cytotoxic and genotoxic responses. J. Biol. Chem. 278: 39951-39959, 2003. PubMed: 12882965

Narayanan L, et al. Elevated levels of mutation in multiple tissues of mice deficient in the DNA mismatch repair gene Pms2. Proc. Natl. Acad. Sci. USA 94: 122-127, 1997. PubMed: 9096356

Sobol RW, et al. Mutations associated with base excision repair deficiency and methylation-induced genotoxic stress. Proc. Natl. Acad. Sci. USA 99: 6860-6865, 2002. PubMed: 11983862

Sobol RW, et al. The lyase activity of the DNA repair protein beta-polymerase protects from DNA-damage-induced cytotoxicity. Nature 405: 807-810, 2000. PubMed: 10866204