Haynes Lab:Notebook/Engineering PC-TFs/2013/10/21

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October 21, 2013

Miniprep liquid cultures

  • Add 100 μL of 7X Lysis Buffer (Blue) to 600 μL of E.coli culture in a 1.5 ml microcentrifuge tube. Mix by inverting tube 4-6times.
  • Add 350 μL of cold Neutralization Buffer (Yellow), mix thoroughly. Neutralization is complete when sample becomes yellow and precipitate has formed.
  • Centrifuge at 11,000-16,000 g for two minutes.
  • Transfer the supernatent into the Zymo-Spin Column. Place column in 2 ml collection tube and centrifuge for 15 seconds at top speed. Discard flow-through.
  • Add 200μL of Endo-Wash Buffer to the column. Centrifuge for 15 seconds at top speed.
  • Add 400μL of Zyppy Wash Buffer to the column. Centrifuge at top speed for 30 seconds.
  • Transfer column to 1.5 microcentrifuge tube and add 50μL of Zyppy Elution Buffer directly to column. Centrifuge for 1 minute at top speed to elute DNA.



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