Daniel:Notebook/ComboLock/2017-7-18

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Multiplex Experiment (Started Thursday July 13)[edit]

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Given yesterday's results, I'm going to repeat the 6 hour RCA and also do a prep heat.

Protocol[edit]

  1. Rolling Circle Amplification
    1. Vortext samples and resuspend with pipette
    2. Heat for 5 minutes at 95C
    3. During this time, prepare master mix according to table below
    4. Reagent Single Rxn (uL) Master Mix X (6.2X)
      Sample 4 24.8
      p12RC*** (10 uM) 0.2 1.24
      RCA Primer LLRC*** (10 uM) 0.2 1.24
      dNTP (10 mM) 0.5 3.1
      10X Buffer 2 12.4
      Phi29 1 0
      nfH2O 12.1 75.02
      Total 20 117.8
    5. Add 19 uL master mix to each tube
    6. Heat reactions up to 95C for 5 min
    7. Cool to 55C and incubate for 15 minutes
    8. Cool to 30C and add 1 uL enzyme
    9. Incubate for 6 hours at 30C
    10. Incubate for 65 minutes at 20C to denature

Continued tomorrow