Daniel:Notebook/HiResChrPaint/2014-1-14

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Alu Control (Started 1/08/2014)

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Dye Coupling Test (with Positive Control)

Repeating the dye coupling test one last time using a positive control as well.

Samples

  • V6S3-A; 13.5 uL, total 1.0 ug
  • Alu2-D; total 1.0 ug
  • Human Cot-1 DNA, 1 ug
0. Vaccuum centrifuge DNA; resuspend in 2.5 uL nfH2O
1. Warm DMSO and bicarbonate labeling buffer and nuclease free water to room temperature
2. Add 1.5 uL of sodium bicarbonate buffer
3. Denature samples for 5 minutes at 95C, then snap cool using ice box
4. Dissolve 1 vial of reactive dye in 2 uL solvent; vortex ≥10 seconds
   *dye cannot be saved for later use.  Use immediately!
5. Add 1 uL dye to sample
6. Incubate in the dark for 1 hour
7. Add 20 uL nfH20 to sample; Measure concentration in Nanodrop
8. Centri-Sep column purification
9. Measure concentration in Nanodrop
Sample ng/uL dsDNA pmol/uL dye pmol/uL ds probe dye/100bp ng/uL ssDNA pmol/uL dye pmol/uL ss probe dye/100bp
Alu2D-Pre Column 170 115     113.2 116.6    
Blank-Pre Column 157.4 111     103.3 110.6    
Cot1-Pre Column 150.9 105     102.1 106.4    
V6S3-Pre Column 160.3 109.2     109.6 112.9    
Alu2D-Post Column 24.4 0.5 0.4 1.4 12.6 0.5 0.4 1.3
Blank-Post Column 1 0 0.0 0.0 0.2 0 0.0 0.0
Cot1-Post Column 25.9 1.7 0.2 13.3 15.6 1.3 0.3 8.4
V6S3-Post Column 41.3 0.6 0.8 0.6 24.4 0.3 0.9 0.3