- Prepare the filter plate for synthesis.
- The filter plate is placed in the vacuum chuck and tightened until there
is no gap left between the two halves of the chuck.
- Using the pipette aid, .003g of universal support CPG's are dispensed into
each of the 96 wells of the filter plate. 500 Å CPG's are used for
oligos up to 40 bases in length. 1000 Å CPG's are used for oligos
over 40 bases in length.
- Setup the machine for synthesis.
- Make sure there are sufficient reagents to complete the run.
- Make sure there is sufficient argon to complete the run.
- Check the waste bottles to make sure they are empty.
- Follow the computer instructions to startup the run.
- Deprotection of the oligos.
- After synthesis, the plate is removed from the machine and its chuck and placed over a 96 well riplate
- 200µl of ammonium hydroxide are added to each well.
- Let stand for 15 minutes and then blow the ammonium hydroxide out of the
synthesis plate into the riplate by placing both plates into the chuck
attached to the compressed air tank. The pressure can range from 5-15psi.
- Repeat with 100µl of ammonium hydroxide for 15 minutes and blow out.
- Cover with a foil sealer and place in baking chucks. Bake at 55º C for 7 hours.
- Allow to cool, remove form baking chucks and dry immediately or store at
4º C overnight. Dry the oligos in the Helix dry down evaporator for 1 hour
and 30 minutes.
- Clean up of the oligos.
- Suspend the dried down oligos in 100µl of nuclease-free water. Vortex
for 2-3 minutes, quick spin.
- Prepare a sephadex G-50 plate and wash the oligos using the Hydra to
transfer form the riplate to the sephadex plate.
- Spin at 1500rpm for 5 minutes.
- Repeat wash and store oligos in freezer.
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