Handling of DNA Oligos

When handling DNA oligonucleotides, please consider the following:

• Make sure you use nuclease-free solutions only (see our suggestion below for potential solvents)
• Handle the DNA oligonucleotides carefully to avoid bacterial contamination
• Oligos are generally speaking more stable at higher concentrations
• Some fluorescent dyes (in particular Cy dyes) are especially light sensitive, and exposure to light should be minimized. Furthermore, some dyes may be sensitive to oxidation. Therefore, please ensure that you only open the tube if required.

How Do You Dissolve Your Oligo?

Generally speaking, oligonucleotides are best dissolved in sterile water or 10 mM Tris-HCl buffer (pH 7.5). However, there are a couple of exceptions that require special attention:

• Oligos carrying a fluorescent dye should not be dissolved in distilled water. This is because distilled water usually has a pH of ~6.0 which favors the degradation of the fluorescent dye. In such cases, you should only use 10 mM Tris-HCl buffer at pH 7.5.
• Oligos carrying fluorescent dyes of the cyanine family, such as Cy3 etc. should not be dissolved in 10 mM Tris-HCl buffer at pH 7.5. This is because they degrade slowly at pH >7.5. In such cases, you should only use distilled water.

How Do You Re-suspend Your Oligo?

• Do a short spin at max. speed in a centrifuge to collect the pellet at the bottom of the tube
• Add an appropriate amount of sterile water or buffer
• Heat the tube for 5 minutes at 65°C
• Vortex or mix by pipetting vigorously up and down

How to Anneal Complementary Oligos?

Option 1:
Materials:
1x annealing buffer (i.e. Tris-HCl 10 mM, pH 7.5-8, 20 mM NaCl or alternative commonly used buffer)

• Dissolve your Oligos at 200 µM in annealing buffer. Therefore, you should use one-half of the volume of the buffer given in the technical datasheet
• Mix the two Oligos in a 1:1 ratio
• Heat the solution to 92°C and keep for 2 min
• Cool to RT by removing the tube from the heating source

Your double strand will be annealed as 100 µM (100 nmol/ml) in Tris-HCl, pH 7.5-8, 20 mM NaCl (or alternative buffer)

Option 2:
Materials:
5x annealing buffer (i.e. Tris-HCl 50 mM, pH 7.5-8, 100 mM NaCl or alternative commonly used 5x buffer)

• Dissolve your Oligo at 100 µM in pure water (information on the volume given is provided in the technical datasheet). Alternatively, you can order the oligos already dissolved as 100 µM (100 nmol/ml) from Microsynth
• Mix the two oligos in a 1:1 ratio
• Add the buffer in a ratio of 4 parts oligo-solution and 1 part 5x annealing buffer 4:1 (v:v)
• Heat the solution to 92°C and keep it at that temperature for 2 min
• Cool to RT by removing the tube from the heating source

Your double strand will be annealed as 40 µM (40 nmol/ml) in Tris-HCl, pH 7.5-8, 20 mM NaCl (or alternative buffer)

Storage of DNA Oligos

If you want to store oligonucleotides, please consider the following:

• Please avoid repeated thawing and freezing as the physical forces involved may degrade your oligos
• Depending on your experiment, please choose a suitable storage condition (we refer you to our suggestions for potential storage conditions set out further below)
• Preparing aliquots may make sense if oligos must be stored and used over a long time period without losing activity