User:Tara K. Luckau/Notebook/Team ConGen/Entry Base

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Tara's Lab Notebook Main project page

Contents

27 September 2010 - Ordered Primers

  • Ordered first set of primers
  • Scun2 and Scun22 (highly variable simple motifs)
    • Lance et al (2009) Conservation Genetics Resources, Sceloporus undulatus
  • Forward and reverse as given in publication, also reverse with pigtail (GTTTCTT) at 5' end, also forward with M13F(-21) tag (TGTAAAACGACGGCCAGT)
  • Try all four combinations
  1. F - R
  2. F - R pigtail
  3. F M13 - R
  4. F M13 - R pigtail
  • If they don't hinder efforts or change optimization decisions, then may make optimization of multiplexes cheaper!

11 October 2010 - PCR Thermalcycler Testing

Purpose

  • 10μL in each well of full-skirt PCR plate, PCR seal
  • First set-up of thermalcycler, established User, Protocols, Plates and Masters
  • Data: 20101011_TKL_CyclerTest.tad

Results

  • Evaporation all around edges
  • Try using silicone mat?

Purpose

  • 10μL in each well of full-skirt PCR plate, PCR seal, 1 silicone mat
  • Data: 20101011_TKL_CyclerTest1Mat.tad

Results

  • Still evaporation around edges, but better
  • Try using 2 silicone mats?

12 October 2010 - PCR Thermalcycler Testing

Purpose

  • 10μL in each well of full-skirt PCR plate, PCR seal, 2 silicone mats
  • Data: 20101012_TKL_CyclerTest2Mat.tad

Results

  • No evaporation!
  • Silicone mats got stuck on lid
  • Try using 2 silicone mats with tape?
  • Try getting samples of different brand of silicone mats, different shape, no holes?

18 October 2010 - PCR Thermalcycler Testing

Purpose

  • 10µL in each well of full-skirt PCR plate, PCR seal, 2 silicone mats, lab tape on opposite sides
  • Data: 20102018_TKL_CyclerTest2MatTape.tad

Results

  • H6, H7 evaporated, but I think it's workable
  • Plan to do first optimization PCR tomorrow!

==18 October 2010 - Primer Rehydration, Scun2

Purpose

  • Rehydrate Scun2 primers for testing tomorrow
  1. Scun2-F
  2. Scun2-R
  3. Scun2-F-M13F(-21)
  4. Scun2-R-pigtail

Calculations


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