Daniel:Protocols/AlexaDyeCouple: Difference between revisions

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ULYSIS Manual from Invitrogen: [[File:ULYSIS_Manual.pdf]]
ULYSIS Manual from Invitrogen: [[File:ULYSIS_Manual.pdf]]


  1.
  1. Add 100 uL DMSO to vial containing ULS dye (stable for 6 months at 4C)
  *For AlexaFluor 488 label using 5 uL DMSO (stable for 1 month at 4C)
2. Resuspend DNA in 20 uL labeling buffer (Component C)
3. Denature DNA at 95C for 5 minutes and snap cool on ice; centrifuge briefly
4. Add appropriate volume of dye to sample (see table 2); if necessary add labeling buffer to bring volume to 25 uL
5. Incubate at 80C for 15 minutes; stop the reaction by plunging the tube into an ice bath
6. Purify using a column (Centri-Sep recommended)
 
[[Image:ULYSIS_ReactionTable.png|thumb|Recommended amount of fluorophore per reaction as well as ratios of dye:base]]

Revision as of 19:10, 3 July 2013

Alexa Fluorophore Dye Coupling

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Dye Couples

Probe Alexa Dye
V4S1 Alexa-546
V4S2 Alexa-488
V4S3 Alexa-594
V4S4 Alexa-647
V6S1 Alexa-546
V6S2 Alexa-488
V6S3 Alexa-594
V6S4 Alexa-647

ARES Protocol

ARES Manual from Invitrogen: File:ARESTM DNA Labeling Kits.pdf

1. Warm DMSO and bicarbonate labeling buffer and nuclease free water to room temperature
2. Add 2.5 uL DNA and 1.5 uL of sodium bicarbonate buffer
3. Denature samples for 5 minutes at 95C, then snap cool using ice box
4. Dissolve 1 vial of reactive dye in 2 uL solvent; vortex ≥10 seconds
   *dye cannot be saved for later use.  Use immediately!
5. Add 1 uL dye to sample
6. Incubate in the dark for 1 hour
7. Add 5 uL 3M NaOAc and 40 uL nfH20 to sample
8. Recommend column purification after incubation

ULYSIS Protocol=

ULYSIS Manual from Invitrogen: File:ULYSIS Manual.pdf

1. Add 100 uL DMSO to vial containing ULS dye (stable for 6 months at 4C)
  *For AlexaFluor 488 label using 5 uL DMSO (stable for 1 month at 4C)
2. Resuspend DNA in 20 uL labeling buffer (Component C)
3. Denature DNA at 95C for 5 minutes and snap cool on ice; centrifuge briefly
4. Add appropriate volume of dye to sample (see table 2); if necessary add labeling buffer to bring volume to 25 uL
5. Incubate at 80C for 15 minutes; stop the reaction by plunging the tube into an ice bath
6. Purify using a column (Centri-Sep recommended)
File:ULYSIS ReactionTable.png
Recommended amount of fluorophore per reaction as well as ratios of dye:base