PCR Cycles

Cyanobacterial Primers (cyano)

First Cycle (run once)

  • 95oC for 5 minutes
  • 80oC for 1 minute
  • With about 3 seconds remaining, pause thermocycler and add Taq

Second Cycle (run 30 times)

  • 94oC for 1 minute
  • 60oC for 1 minute
  • 72oC for 1 minute

Third Cycle (run once)

  • 72oC for 9 minutes
  • 4oC hold

Eubacterial Primers (eubac td)

First Cycle (run once)

  • 95oC for 5 minutes
  • 80oC for 1 minute
  • With about 3 seconds remaining, pause thermocycler and add Taq
  • 65oC for 1 minute
  • 75oC for 3 minutes

Second Cycle, Touchdown PCR (run 20 times)

  • 94oC for 1 minute
  • 65oC for 1 minute
  • 75oC for 3 minute
  • NOTE: this is a touchdown cycle; the temperature for the second step must be lowered by 1oC every second cycle (ending on 55oC).

Third Cycle (run 9 times)

  • 94oC for 1 minute
  • 55oC for 1 minute
  • 72oC for 3 minute

Fourth Cycle (run once)

  • 72oC for 7 minutes
  • 4oC hold