Daniel:Notebook/HiResChrPaint/2013-7-15: Difference between revisions
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>Djacobse (Created page with "=S2 Probe Prep (Started 5/29/2013)= Back to Calendar ==ARES Labeling of V6S2== Trying ARES lab...") |
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Trying ARES labeling of [[Daniel:Notebook/HiResChrPaint/2013-6-10#Nanodrop Results|V6S2 set]] randomly primed with aa-dUTP. | Trying ARES labeling of [[Daniel:Notebook/HiResChrPaint/2013-6-10#Nanodrop Results|V6S2 set]] randomly primed with aa-dUTP. | ||
'''V6S2 ~ | '''V6S2 ~ 1.2 ug DNA''' | ||
*Following [[Daniel:Protocols/AlexaDyeCouple#ARES|ARES Protocol | *Following [[Daniel:Protocols/AlexaDyeCouple#ARES|ARES Protocol]] | ||
1. Warm DMSO and bicarbonate labeling buffer and nuclease free water to room temperature | 1. Warm DMSO and bicarbonate labeling buffer and nuclease free water to room temperature | ||
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6. Add 2 uL dye to sample | 6. Add 2 uL dye to sample | ||
7. Incubate in the dark for 1 hour | 7. Incubate in the dark for 1 hour | ||
8. Add | 8. Add 15 uL nfH20 to sample | ||
9. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column | 9. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column | ||
==ULYSIS Labeling of V6S2== | ==ULYSIS Labeling of V6S2== | ||
Trying ULS labeling of [[Daniel:Notebook/HiResChrPaint/2013-6-10#Nanodrop Results|V6S2 set]] randomly primed with aa-dUTP. | |||
'''V6S2 ~ 1.5 ug DNA''' | |||
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; AmpR13: Dye from 7/10/13; AmpR14: Dye from 7/11/13 | |||
4. Incubate for 20 minutes at 80C, snap cool on ice | |||
5. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column |
Revision as of 17:54, 15 July 2013
S2 Probe Prep (Started 5/29/2013)
ARES Labeling of V6S2
Trying ARES labeling of V6S2 set randomly primed with aa-dUTP.
V6S2 ~ 1.2 ug DNA
- Following ARES Protocol
1. Warm DMSO and bicarbonate labeling buffer and nuclease free water to room temperature 2. Resuspend DNA in 2.5uL nfH20 3. Add 5 uL DNA and 3 uL of sodium bicarbonate buffer in PCR tube 4. Denature samples for 5 minutes at 95C, then snap cool using ice box 5. Dissolve 1 vial of reactive dye in 2 uL solvent; vortex ≥10 seconds *dye cannot be saved for later use. Use immediately! 6. Add 2 uL dye to sample 7. Incubate in the dark for 1 hour 8. Add 15 uL nfH20 to sample 9. Purify using Centri Sep column
ULYSIS Labeling of V6S2
Trying ULS labeling of V6S2 set randomly primed with aa-dUTP.
V6S2 ~ 1.5 ug DNA
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; AmpR13: Dye from 7/10/13; AmpR14: Dye from 7/11/13 4. Incubate for 20 minutes at 80C, snap cool on ice 5. Purify using Centri Sep column