Daniel:Notebook/ComboLock/2017-4-5

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RCA Test (Started March 24)

Back to Calendar

Second Strand Synthesis part II

Since the second strand synthesis from Monday worked well, I'm going to repeat the experiment but using the NEB 3 hr and 6 hr samples (X and P, respectively). I won't bother with the Bgl digested ones as I'll just digest after.

  1. qPCR-Second strand synthesis
    1. Make the following 14.2X master mix
      1. 14.2 uL AmpF-6.4Sol (10 uM)
      2. 284 uL Kapa SYBR 2X Master Mix
      3. 241.4 uL nfH2O
    2. Aliquot 38 uL master mix into each lane
    3. Add 2 uL sample according to plate layout
    4. File:PlateLayout-qPCR-20170405-RCATest-CPLP-ssSynthesis2.png
    5. Run the following protocol
      1. 95C for 2 min
      2. 95C for 5 sec
      3. 55C for 30 sec
      4. 72C for 45 sec
      5. Goto (b) x20
      6. 72C for 2 min