Haynes Lab:Notebook/Engineering PC-TFs/2014/03/12

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Summary

02/14/14

  • Gel Purification: CMV/V0120, MV9



Using Prior Miniprep Digests

  1. CMV culture 1 - check with E/P
  2. MV9 culture 2 - check with P/X
  • Used previously digested CMV and MV9.
  • Placed 15μL 1kB Ladder in well one and 30μL of both CMV and MV9 digest in well two and three respectively.
  • Ran gel at 110V for 30 minutes.
  • Placed gel on UV transilluminator and cut out desired bands.
Hover name
15 μL/lane; 1% agarose; Ladder
  • Used Zymo gel purification kit.
  • Assumed 200mg gel, added 600μL ADB to each 1.5mL tube.
  • Put on heat block at 55°C for 5 minutes, vortexed and then placed back on the heat block for an additional 5 minutes, vortexed again.
  • Put ~750μL gel/ADB into spin column, centrifuged at max rpm for 30 seconds.
  • Pipetted 200μL wash buffer to each tube, centrifuged at max rpm for 30 seconds. Repeated once.
  • Transferred columns to new labeled 1.5mL tubes.
  • Pipetted 15μL elution buffer to each column.
  • Spun at max rpm for 30 seconds.
  • Put marked CMV and MV9 in -20°C fridge in box labeled Cameron.







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