PdaeMap/30-Mar-2016

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Update: 09:45 Mar 30, 2016 By: EliMeyer

Today we did test scale PCR

Component 1x 11x
dNTP 10 mM 0.5 5.5
ILL-Lib1, 10 µM 0.5 5.5
ILL-Lib2, 10 µM 0.5 5.5
ILL-HT, 1 µM 1.3 13.8
ILL-BC, 1 µM 1.3 13.8
BSA 20 mg / ml 0.3 2.8
Q5 buffer, 5x 5.0 55.0
Q5 Taq polymerase 0.3 2.8
NFW 5.5 60.5

great success for my samples! partial success for Emily's.

2016 03 30 2697.b.JPG

Update: 15:12 Mar 30, 2016 By: EliMeyer

Did additional pooled larval extractions to test RNALater vs EtOH for larval preps

  1. used Omega Biotek DNA kit
  2. ~10 larvae per extraction
  3. started with 200 µl TL, 25 µl OB protease, 0.5 µl RNAse
  4. minimal lysis at 55°C
  5. eluted in 2x 35 µl, Vt = 35 µl
  6. quantified by Accuclear

terrible standard curve (r2 = 0.89) but I estimate these concs

sample ng / µl
EtOH 32
RNALater 30