Friday 10/23/15

9:00 am

pairing meeting

  • rehearse talk (8:45-9:15)
  • see notes

Experiments: QPCR for RNAi (9:15 am – 11:05 am)

  • RNA concentrations from yesterday: 600 – 1200 ng/uL (55 uL volume)
  • ran RNase H digestion last night, at 4C in block O/N.
  • ran clean up column (DCC5)
  • cDNA concentrations from polyT primed RT reaction: 40 – 100 ng/uL

Setting up QPCR

  • 40x master mix (9 primer pairs, 4 samples)
  • per each (total for master)
    • 12.5 uL Hot-start Phusion mix (500 uL)
    • 2.5 uL Eva Green (50 uL)
    • 6 uL ddH2O (240 uL)
    • 2 uL cDNA
    • 2 uL primer
  • plate layout
    • col 1:11 = Act, Tbu, Gapdh, ubx, Abd-B, Dfd, Antp, en, ph-p, ph-d1, ph-d2,
    • rows 1:4 = PhWT 1 10-16, PhWT 2 10-16, Wt 2 day 10-22, Ph 2 day 10-22.
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