| 1. |
Determine the concentration of your oligonucleotide solutions. Dissolve
lyophilized oligonucleotide in water or TE and verify concentration by
determining the A260 value of duplicate dilutions. Use the conversion factor
printed on the oligonucleotide label to determine nanomole/A260 unit
|
| 2. |
Add the following components together:
|
| Stock |
Final Concentration |
| Oligonucleotide
1 |
100 nanomoles/milliliter |
| Oligonucleotide
2 |
100 nanomoles/milliliter |
| * 10X
Annealing Buffer |
1X |
| Nuclease-free
water |
to
appropriate volume |
* 10X Annealing Buffer: 100 mM Tris-HCl, pH 7.5, 1 M NaCl, 10 mM EDTA.
|
3. |
Heat the oligo solution to a temperature 10° C higher than the calculated melting
temperature. Maintain the temperature for 10 minutes. Remove the solution from
the heating block/water bath and allow it to cool slowly to room temperature on
the bench (approximately 1 hour).
|
4. |
Store the annealed oligos at -20° C.
|