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

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  5. Incubate for 15 minutes at 80C, snap cool on ice
  5. Incubate for 15 minutes at 80C, snap cool on ice
  6. Purify using [[Daniel:Protocols/CentriSep|Centri-Sep Column]]
  6. Purify using [[Daniel:Protocols/CentriSep|Centri-Sep Column]]
===Results===
{| class="wikitable" <hiddentext>generated with [[:de:Wikipedia:Helferlein/VBA-Macro for EXCEL tableconversion]] V1.8</hiddentext>
|- style="font-size:12pt" align="center"
| align="center" width="65" height="30" | &nbsp;
|style="font-weight:bold" width="65" | ng/uL DNA
|style="font-weight:bold" width="65" | pmol/uL dye
|style="font-weight:bold" width="65" | ug DNA in 20 uL
|style="font-weight:bold" width="65" | pmol/uL DNA
|style="font-weight:bold" width="65" | dye/probe
|style="font-weight:bold" width="65" | dye/100 bp
|style="font-weight:bold" width="65" | Base:Dye Ratio (:1)
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | AmpR4
| align="center" | 31.8
| align="center" | 1.4
| align="center" | 0.636
| align="center" | 1.9
| align="center" | 0.7
| align="center" | 1.45
| align="center" | 69
|- style="font-size:12pt" align="center"
|style="font-weight:bold" height="15" | BLANK
| align="center" | 66.6
| align="center" | 0.2
| align="center" | 1.332
| align="center" | 4.0
| align="center" | 0.0
| align="center" | 0.10
| align="center" | 1009
|}

Revision as of 18:53, 9 July 2013

S2 Probe Prep (Started 5/29/2013)

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ULYSIS Dye Coupling

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