Daniel:Notebook/HiResChrPaint/2013-7-9: Difference between revisions

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[[Daniel:Notebook/HiResChrPaint|Back to Calendar]]
[[Daniel:Notebook/HiResChrPaint|Back to Calendar]]


==ULYSIS Dye Coupling==
==ULYSIS Dye Coupling (Test 3)==


  Based on [[Daniel:Notebook/HiResChrPaint/2013-7-8|yesterday's results]] I think I'm going to see what happens if I use all 5 uL of dye available (see ULYSIS protocol)
  Based on [[Daniel:Notebook/HiResChrPaint/2013-7-8|yesterday's results]] I think I'm going to see what happens if I use all 5 uL of dye available (see ULYSIS protocol)

Revision as of 20:21, 9 July 2013

S2 Probe Prep (Started 5/29/2013)

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ULYSIS Dye Coupling (Test 3)

Based on yesterday's results I think I'm going to see what happens if I use all 5 uL of dye available (see ULYSIS protocol)
in one labeling. I'll use the oligo Noi gave me again, as I have a lot and it doesn't waste probe.
1. Add 5 uL DMSO to Alexa dye 488
2. Add 1 uL DNA in 19 uL labeling buffer (Component C)
3. Denature DNA for 5 minutes at 95C; snap cool on ice
4. Add 5 uL dye to sample
5. Incubate for 15 minutes at 80C, snap cool on ice
6. 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)
AmpR4 31.8 1.4 0.636 1.9 0.7 1.45 69
BLANK 66.6 0.2 1.332 4.0 0.0 0.10 1009
This is not far off from the suggested efficiency values (44-67:1).  I think that the efficiency can be improved, however, 
by also incubating longer.