Daniel:Notebook/HiResChrPaint/2013-7-11

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Revision as of 19:46, 11 July 2013 by >Djacobse
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S2 Probe Prep (Started 5/29/2013)

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ULYSIS Dye Coupling (Test 6)

Should be the last test.  I'm going to test at 15 minutes and 20 minutes incubation time, 1 uL each.  If the
results go well this morning I will be labeling the V6S2 set this afternoon. 
1. Add 1 uL DNA in 19 uL labeling buffer (Component C)
2. Denature DNA for 5 minutes at 95C; snap cool on ice
3. Add 1 uL dye to sample; add 4 uL labeling buffer
4. Incubate for 15 minutes (AmpR9) or 20 minutes (AmpR10) at 80C, snap cool on ice
5. Purify using Centri-Sep Column

Results

  ng/uL DNA pmol/uL dye ug DNA in 20 uL pmol/uL DNA dye/probe dye/100 bp Base:Dye Ratio (:1)
AmpR9 64.6 0.6 1.3 3.9 0.2 0.3 326
AmpR10 71.1 0.4 1.4 4.3 0.1 0.2 539
It would seem that I can always tell beforehand whether a yield will be good or bad, by the color of
the solution post incubation.  If the solution has a faint green color it will be a good yield.  If 
the solution remains clear it will be a bad yield.  Therefore I'm going to ignore columns for future 
trials (yes, it looks like I need more troubleshooting).  That will save on time and $$, and hopefully
I'll get to the bottom of this before long.