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DNA Preparation for PCR Product Less Than 500 Bp

Lysis Buffer

 
Final Concentration
Stock Concentration
For 10 ml Mix
Tris pH8.8 50 mM 1 M 500 l
EDTA pH 8.0 1 mM 0.5 M 20 l
Tween 0.5% 10% 500 l
H2O     9000 l

  • Add Proteinase K before use.
  • For tail piece 0.2 mm long or ear piece, add 50 ul buffer plus 1.5 ul of 20 mg/ml (30 ug) Proteinase K stock solution.
  • Ten samples: 550 ul lysis buffer + 16.5 ul PK 20 mg/ml.
  • You can use 0.5 ml PCR tubes for digestion and heat in a PCR machine.

Lysis

  • 55C, O/N , then heat inactive at 100C for ten minutes. Cool down on bench top.
  • For 20 ul PCR reaction, use 0.8 ul tail DNA.