NIH3T3/3790#30 rho-0 cell line

NIH3T3 cell line lacking mitochondrial DNA and oxidative phosphorylation

Contributors

Inventor Institute
Mikhail Alexeyev University of South Alabama Foundation for Research and Commercialization
Cat. #: 162192
Organism: Mouse
Gender: Male
Model: Genetically modified cell line
Morphology: Epithelial
Growth properties: Adherent
Primary citation: Spadafora et al. 2016. PLoS One. PMID: 27136098
Product description: NIH3T3 cell line lacking mitochondrial DNA and oxidative phosphorylation. Auxotrophic for uridine and pyruvate. Can be used for generation of transmitochondrial cybrids, control in experiments studying mtDNA contribution to tumor initiation, promotion, malignant conversion, and progression.
Gender: Male
CRISPR: No
Conditional: No
Production details: Prolonged incubation with an inhibitor of mtDNA replication followed by cloning
Additional notes: Cells rely exclusively on glycolysis for their growth. Therefore, they tend to consume glucose faster than their WT counterparts, acidify the medium faster, and die as soon as they consume all glucose. Therefore, do not allow excessive medium acidification (an indicator of glucose depletion) and do not grow cells to high densities
Parental cell line: NIH3T3
Model description: Lacks mitochondrial DNA and all genes encoded there
disease: Mitochondrial Disease
Initial handling information: Upon arrival, store the vial in liquid nitrogen. Quickly thaw the vial in a 37C water bath, transfer vial's content into a 15-ml centrifuge tube containing complete growth medium, pellet cells by centrifugation, remove the medium, resuspend the cells in 10 ml of prewarmed complete growth medium, transfer to 100-mm TC-treated petri dish, incubate @37C+5% CO2.
Format: Frozen
Storage conditions: Liquid Nitrogen
Shipping conditions: Dry ice
Growth medium: DMEM medium with 4.5 g/l glucose supplemented with 10% FBS, 50 ug/ml gentamycin, 50 ug/ml uridine and 1 mM sodium pyruvate
Subculture routine: Do not grow to confluence and do not allow medium acidification. When the growth medium starts changing its color, immediately change the medium or passage cells. To passage, aspirate growth medium, wash with PBS (without Ca++ or Mg++), aspirate PBS, add 0.05% Trypsin in PBS, incubate @37C, 5% CO2 for 5-10 min, tap the vessel to dislodge cells, neutralize trypsin by adding 5 volumes of complete growth medium, disperse cells by trituration, plate 1:4 to 1:10 dilution
Atmosphere: 5% CO2
Storage medium: (DMEM medium with 4.5 g/l glucose supplemented with 10% FBS, 50 ug/ml gentamicin, 50 ug/ml uridine and 1 mM sodium pyruvate)=45% + (Heat-inactivated newborn calf serum)=45% + 10% sterile DMSO
Recommended controls: WT NIH3T3 cells
References:

Spadafora et al. 2016. PLoS One. PMID: 27136099

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