User:Torsten Waldminghaus/Notebook/dnaC2 temperature shift experiment

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Principle

The dnaC2 mutation in E. coli can be used as tool to synchronize DNA replication. The DnaC protein is needed for initiation of replication. Temperal inactivation via temperature shift of the ts mutant DnaC2 leeds to stop of initiation. Shift back to lower temperature leads to restart of replication.

Protocol

  • Grow 100ml culture in desired medium at 30°C starting the culture with an over night culture 1:1000
  • At OD 0.075 shift the culture to 39°C
  • after 30 min put the flask back to 30°C and add
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