Team:Tokyo Metropolitan/Notebook/Fiber/2010/08/12

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E.coli Fiber Project Notebook

August 2010
SUNMONTUEWEDTHUFRISAT
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September 2010
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October 2010
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2010/08/12 Tuesday (Bambi75)

Experiment:PCR

Member
NEX , bambi75 and watachin
Materials
  • sterilized water 106.5μl
  • Ex taq buffer 15μl
  • dNTP 12μl
  • Ex taq 1.5μl
  • K12bcsA sense(10μmol/l)2.5μl
  • K12bcsA antisense(10μmol/l)2.5μl
  • K12bcsB sense(10μmol/l)2.5μl
  • K12bcsB antisense(10μmol/l)2.5μl
  • K12bcsC sense(10μmol/l)2.5μl
  • K12bcsC antisense(10μmol/l)2.5μl
  • E.coliK12strain
Equipment
  • thermal cycler
  • vortex mixer
  • PCR-tubes
  • pipette tip
Procedure
1.mix materials : sterilized water , Ex taq buffer , dNTP and Ex taq and divide 1) into 3 tips.
2.*mix K12bcsAsense , K12bcsAantisense and 2)
mix K12bcsBsense , K12bcsBantisense and 2)
mix K12bcsCsense , K12bcsCantisense and 2)
3.dip E.coliK12strain into 3).
4.setting tips and elongation in thermal cycler
  • initialization 95℃ 3min
  • denaturation 96℃1min
  • annealing 55℃ 1min
  • elongation 72℃ 5min (30 cycles from 95℃ to 72℃)
  • reaction stop 10℃