|
|
 |
|
| Home
> Microarrays > Protocols > Labeling Amino-linked Oligos |
|
|
Note: make sure that the amino-labeled oligos are resuspended in dH2O not TE (Tris will compete the labeling reaction).
- Dry down 5,000 pmols of amino-labeled oligo (either the same oligo or a mixture of oligos) in a speed vac
- Resuspend in 30 microliters 2X Fluor labeling buffer (Clontech)
- Resuspend 1 Cy3 or Cy5 dye pack (Amersham cat. # PA23001 or PA25001) in 45 microliters high quality DMSO (this solution may be stored in the dark at -20° C for several months)
- Add 30 microliters Cy dye-DMSO solution to oligo mixture
- Incubate in the dark at room temp for 30 min.
- Add 6 microliters 3M sodium acetate (pH 5.2) & mix well
- Add 150 microliters 100% ethanol & mix well
- Freeze at -20° C for > 1 hour (overnight OK)
- Centrifuge 20 min. (eppendorf cfg. full speed) @ 4° C
- Pipet off supernatant
- Add 200 microliters 70% ethanol & mix well
- Centrifuge 5 min. (eppendorf cfg. full speed) @ 4° C
- Pipet off supernatant
- Resuspend in 400 microliters dH2O
- Concentrate through a microcon-10 (Millipore cat. # 42406) filter to desired volume
(do not speed vac)
Information on quantitating labeling efficiency can be found in the Clontech Atlas Glass Fluorescent Labeling Kit Manual.
Other protocols:
- Staining Slides with Sybr Green HTML | PDF
- cDNA Synthesis and Labeling HTML | PDF
- Labeling Amino-linked Oligos HTML | PDF
- Pre-hybridizing Polylysine Slides HTML | PDF
- Pre-hybridizing CodeLink Slides HTML | PDF
- Hybridization and Washing HTML | PDF
|
|
|
