Daniel:Notebook/PosSequencing/2017-6-2

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Emulsion PCR Test 3 (Started Wednesday)

Back to Calendar

Protocol-Part 2

  1. Single Primer qPCR
    1. Make the following master mixes
    2. Reagent uL in Single Rxn Master Mix P2 (5.2X) Master Mix BioU (5.2X) Master Mix Dual (5.2X)
      Primer 2 (10 uM) 1 5.2 0 5.2
      BioU Primer (10 uM) 1 0 5.2 5.2
      2X Kapa SYBR Master Mix 20 104 104 104
      nfH2O 17 88.4 88.4 83.2
      Total* (+ 2 uL sample=40) 38 197.6 197.6 197.6
    3. Aliquot 38 uL master mix to appropriate lanes
    4. Add 2 uL sample according to plate layout below
    5. File:PlateLayout-20170601-SPqPCR-emPCRTest3-Round2.png
    6. Run the following thermocycler program
      1. 95C 3 min
      2. 95C 3 sec
      3. 55C 30 sec
      4. 72C 20 sec
      5. plate read
      6. goto b x150
      7. 72C 2 min
      8. 16C hold