Daniel:Notebook/HiResChrPaint/2013-5-22

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Probe Prep Test 2 (Started 5/10/2013)[edit]

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Dye Coupling (Round 2)[edit]

After talking with Dr. Zhang yesterday, he suggested I try the dye coupling with just 1 reaction, and purify with a standard column purification.

Protocol[edit]

1. Warm DMSO, Sodium bicarbonate, and V4S3 in 37 degree shaker
2. Add 2.5 uL nf H20 to dna (should bring total to ~5)
3. Denature at 95C for 5 minutes
4. Add 3 uL sodium bicarb to reaction
5. Add 2 uL DMSO to dye, vortex ≥10 seconds
6. Add 2 uL dye/DMSO to reaction; vortex ≥10 seconds
7. Incubate in the dark for 1 hour
8. Add 40 uL nf H20
9. Column purification, add 10x PB, elute with 30 uL nfH20
10. Measure in nanodrop using microarray setting

Nanodrop Results[edit]

ng/uL DNA pmol/uL dye pmol/uL DNA dye/probe ug dye Yield (%)
16.5 2.2 0.62 3.564 0.495 11.0

Discussion[edit]