Daniel:Notebook/HiResChrPaint/2013-7-15: Difference between revisions

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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 15 uL nfH20 to sample
  8. Add 11 uL nfH20 to sample
  9. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column
  9. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column


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*Following [[Daniel:Protocols/AlexaDyeCouple#ULYSIS|ULYSIS Protocol]]
*Following [[Daniel:Protocols/AlexaDyeCouple#ULYSIS|ULYSIS Protocol]]


  1. Add 1 uL DNA in 19 uL labeling buffer (Component C)
  1. Resuspend DNA in 20 uL labeling buffer (Component C)
  2. Denature DNA for 5 minutes at 95C; snap cool on ice
  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
  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
  4. Incubate for 20 minutes at 80C, snap cool on ice
  5. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column
  5. Purify using [[Daniel:Protocols/CentriSep|Centri Sep]] column

Revision as of 19:27, 15 July 2013

S2 Probe Prep (Started 5/29/2013)

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ARES Labeling of V6S2

Trying ARES labeling of V6S2 set randomly primed with aa-dUTP.

V6S2 ~ 1.2 ug DNA

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 11 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. Resuspend DNA in 20 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