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Gitschier Protocol for Genomic DNA isolation from mice

10x Modified Gitschier Buffer (10x MGB)
223 mL         Tris (1.5 M, pH 8.8)= 670 mM Tris
10.97 g         Ammonium Sulfate =166 mM Ammonium Sulfate
3 g                Magnesium Chloride= 65 mM Magnesium Chloride
Fill to 500 mL with Milli-Q H20, mix well and autoclave

1x MGB (for 10 mL)
8.85 mL        ddH2O
1 mL             10x MGB
100 μL          2-mercaptoethanol
50 μL            Triton X-100
Vortex vigorously to make it soluble Triton X-100

Proteinase K mastermix
10 parts         1x MGB
1 part             20 mg/mL Proteinase K

Method:
  1. Clip 5mm mouse tail with a clean razor blade, place in 1.5 Ll tube, and store at -20 ̊C until further processing.
  2. Add 100 μL of 1xMGB
  3. Heat at 95 ̊C for 10 minutes
  4. Add 110 μL of Proteinase K mastermix (10 parts 1x MGB, 1 part 20 mg/mL Proteinase K)
  5. Incubate at 55 ̊C for 30 minutes
  6. Vortex vigorously for 1 second
  7. Repeat steps 5-6 two more times
  8. Spin tubes at 14,000 x g for 5 seconds
  9. Heat at 95 ̊C for 10 minutes
  10. Spin at 14,000 x g for 5 minutes. Store at -20 ̊C until use. Use 1 μL per 50 μL PCR reaction.