CELL CULTURE OF MOUSE MYOBLASTS |
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Last Update: December 2006 |
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Based on the following publication: Delgado I, Huang X, Jones S, Zhang L, Hatcher R, Gao B, Zhang P (2003) Dynamic gene expression during the onset of myoblast differentiation in vitro. Genomics 82: 109-121 |
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| PREPARE SOLUTIONS | |
| 1. Dulbecco's Modified Eagle Medium (DMEM) (500 mL): | Purchase from Invitrogen (11971-025). Contains 4,500 mg/L D-glucose, and L-glutamine, but no sodium phosphate or sodium pyruvate. Store at 4oC |
| 2. Fetal Bovine Serum (500 mL): | Purchase from Invitrogen (10437-028). Store at -20oC |
| 3. Basic fibroblast growth factor (bFGF) (10 mg): | Purchase from Invitrogen (13256-029) (also known as Heparin Binding Growth Factor (HBGF)-2) (Human, Recombinant). Store at -20oC |
| 4. Horse serum (100 mL): | Purchase from Invitrogen (16050-130). Store at -20oC |
| PROCEDURE | |
| 1. Obtain C2C12 mouse myoblasts from the American Type Culture Collection (ATCC, Manssas, VA) | |
| 2. Maintain the myoblasts in DMEM medium supplemented with 20% Fetal Bovine Serum (FBS) | |
| 3. Let the cells grow to 80% confluency prior to inducting muscle differentiation | |
4. Induce cells to differentiate by adding differentiation medium (DMEM supplemented with 2% horse serum) |
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| NOTE: add basic fibroblast growth factor (bFGF, Sigma, Saint Louis, MS) to the differentiation medium to a concentration of 25 ng/mL to inhibit myoblast differentiation | |
| 5. Cells can be collected before and after differentiation and at any time internal of interest | |