RNA oligos are susceptible to degradation by exogenous ribonucleases introduced during handling. Your RNA oligos should not be handled with ungloved hands. RNase-free reagents and supplies should be used. Dried RNA oligos are stable for 1 year at -20C.
Concentrations
To make a 100uM (umole/L or pmole/uL) solution: nanomole * 10 = volume in uL to re-suspend.
Example: adding to 200uL of media to 20 nanomoles of oligo would yield 100uM of solution.
Re-suspension of Annealed siRNA Duplex
Spin down the tube containing siRNA duplex to collect the contents at the bottom of the tube. Add DEPC water to the annealed siRNA duplex and heat it to 90ºC for 1 minute. Then incubate at 37ºC for 60 minutes.
All Retrogen siRNA duplexes are provided purified, lyophilized, and ready for use upon re-suspension. Once re-suspended, oligonucleotide stock solutions are best kept frozen at -20ºC. They may be stored in this state for several weeks and may remain stable for several months. The most important factor in storing working solutions is to use nuclease-free, sterile water. Drying down of your oligos and keeping them at -20ºC is recommended for long-term storage.
Annealing of RNA Oligos
In an RNase-free micro-centrifuge tube, combine the sense and antisense RNA oligonucleotides, water, and 5X RNA Annealing Buffer. The final concentration should be 20 – 100uM for each oligo in 1X Annealing Buffer. Heat this for 1 min at 90ºC followed by incubation for 1 hr at 37ºC. Once annealed, the double stranded RNA is more nuclease resistant and can be stored at -20ºC in a non-frost-free freezer.
Annealing Buffer:
30mM HEPES (pH 7.4)
100mM Potassium Acetate
2mM Magnesium Acetate
This solution can be stored frozen at -20ºC and refrozen many times.
1-800-RETROGEN
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