Yu:quick miniprep

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Contents

Procedure

  1. Spin down 1-3 mL culture
    1. 4000 RPM for 5min at 4°C
  2. Transfer pellet to eppendorf tube if not already in one
  3. Resuspend pellet in 200 μL P1
  4. Add 200 μL of P2
    1. Mix by inversion - solution will become very viscous
  5. Add 200 μL of P3
    1. Mix by inversion - white precipitate will form
  6. Spin mixture for 10 min at top speed
  7. Remove pellet with a sterile toothpick
  8. Add 600 μL Isopropanol
  9. Spin mixture for 3 min at top speed
    1. Decant supernatant
  10. Add 500 µl 70% EtOH and vortex briefly
  11. Spin mixture for 3 min at top speed
    1. Decant supernatant
  12. Air dry pellet
  13. Resuspend pellet if 50-100 μL TE8

Solutions

P1

Final Concentration For 50 mL For 100 mL
50 mM Tris pH8.0 2.5 mL of 1M Tris pH8 5 mL of 1M Tris pH8
20 mM EDTA 2.0 mL 0f 0.5 M EDTA 4.0 mL 0f 0.5 M EDTA
milliQ H2O to 50 mL to 100 mL


P2

Final Concentration For 50 mL For 100 mL
0.2N NaOH 1 mL of 10 N NaOH 2 mL of 10 N NaOH
1% SDS 2.5 mL 0f 20% SDS 5 mL 0f 20% SDS
milliQ H2O to 50 mL to 100 mL

P3

3M NaOAc pH4.8

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