Daniel:Notebook/ComboLock/2017-2-2

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Revision as of 17:47, 2 February 2017 by >Djacobse (→‎Results)
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RCA Time Test (Started Friday Jan 27)

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Phosphorothiorate RCA Primer (Started Yesterday)

  1. qPCR
    1. Make qPCR master mix according to following recipe
      1. 361.2 uL nfH2O
      2. 17.2 uL AmpF6.4
      3. 430 uL Kapa SYBR Fast
    2. Add 47 uL master mix to each well
    3. Add 2 uL sample and 1 uL AmpR indexed according to table
    4. Sample Sample Condition AmpR Index Lane Sample Vol (uL) 2X Kapa SYBR qPCR MM 10 uM Forward Primer 10 uM Reverse Primer H2O Total Volume (uL)
      Sample 1AN Sample 3 hours 20 A8 2 25 1 1 21 50
      Sample 1BN Sample 3 hours 20 A7 2 25 1 1 21 50
      Sample 3AN No C Probe 3 hours 21 A6 2 25 1 1 21 50
      Sample 3BN No C Probe 3 hours 21 A5 2 25 1 1 21 50
      Sample 1AO Sample 3 hours-Epi 22 A4 2 25 1 1 21 50
      Sample 1BO Sample 3 hours-Epi 22 A3 2 25 1 1 21 50
      Sample 3AO No C Probe 3 hours-Epi 23 A2 2 25 1 1 21 50
      Sample 3BO No C Probe 3 hours-Epi 23 A1 2 25 1 1 21 50
      NTC     24 H1 0 25 1 1 23 50
    5. qPCR Cycles
      1. 95C 3 min
      2. 95C 3 sec
      3. 55C 30 sec
      4. 72C 20 sec
      5. plate read
      6. goto b x24
      7. 72C 2 min
      8. 16C hold
  2. TBE Gel
    1. Mix 94 uL TBE, 24 uL 6x loading dye
    2. Aliquot 10 uL per sample/ladder lane onto parafilm
    3. Add 2 uL of sample or ladder to correct drop
    4. Load 10 uL in to well
    5. Run gel for 23 minutes at 230V
    6. Open gel and stain with 3 uL SYBR Gold for 3 minutes
    7. Rinse gel and image in gel doc

Results