Protocol for Annealing Oligonucleotides
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1. Prepare a solution of 10X annealing buffer.
10 mM MgCl2 , 200 mM Tris-HCl, pH 8.0
2. Prepare an aliquot of the oligonucleotide probe in an eppendorf tube.
3. Add a molar excess of oligonucleotide target to the tube.
4. Calculate the volume of 10X annealing buffer to add.
Reaction Volume = Volume of Probe + Volume of Target
Volume of 10X Annealing Buffer = 9 x Reaction Volume
5. Add the 10 X Annealing buffer and vortex to mix.
6. Place the tube in a 95o C Heat Block for 3 minutes.
7. Remove the tube and let it cool to room temperature. (~10 minutes)
8. Vortex the mixture
