Using Minor Bases on an Expedite Synthesizer

The Expedite 8900 series synthesizers are excellent DNA synthesizers, which use very small amounts of amidites per coupling cycle. However, the volume of reagent needed to purge, dry and prime the lines and valve block can require multiple priming steps when changing an amidite. This is a significant disadvantage, especially when using an expensive amidite. There are, however, a few measures that can be taken to decrease the volume used by the Expedite 8900 series synthesizer.

  1. Put 5ml of anhydrous acetonitrile in a clean dry amidite vial and put this vial on the synthesizer in the minor base position that is to be used.
  2. In the "Tools"-"Diagnostic"-"Fluids" mode, using column #1, flush lines with the ACN, first using the "flow" command, then using the volume command. Repeat this process for column #2. This will wash the lines and valve block with anhydrous acetonitrile and help scavenge any moisture present in the lines.
  3. Repeat as necessary until the vial is empty.
  4. Repeat the "flow" command 3-5 times to flush ACN from the frit and lines. This will dry out the lines with helium so that the amidite loaded will not be diluted.
  5. Remove the empty vial and wipe the minor base line and frit with a Kimwipe.
  6. Put the vial containing the amidite to be used on the synthesizer in the minor base position and prime using either the "Prime All" or "Prime Monomers" command. These prime procedures use 10 pulses to prime each of the amidites. This will consume approximately 160µl of amidite. If you have excess amidite you might want to repeat the "Prime Monomers" to prime with an additional 160µl of amidite. Note the "Prime Individual" command uses 20 pulses (320µl) to prime each reagent selected. Experienced users can use the "prime individual" and stop the cycle after counting 10 pulses.

Amidite Consumption:

The Expedite 8905 and 8909 DNA synthesizer delivers reagents in pulses. Each pulse delivers approximately 16µL of solution.
Step # pulses Volume delivered (µL)
Prime 10 160
Amidite (0.2 µmole) 5 80
Amidite (1 µmole) 7 112

Maximum number of couplings per pack
(Approximate dependingon actual flow rates)

Pack size Dilution Vol. Syn. Scale # Primes # Couplings Vol. Used
50 µmole 0.5 ml 0.2 µmole 1 4 480 µL
2 2 480 µL
1.0 µmole 1 3 498 µL
2 1 432 µL
50 µmole 0.75 ml 0.2 µmole 1 7 720 µL
2 5 720 µL
1.0 µmole 1 5 720 µL
2 3 656 µL
100 µmole 1.0 ml 0.2 µmole 1 10 960 µL
2 8 960 µL
1.0 µmole 1 7 944 µL
2 6 992µL
100 µmole 1.5 ml 0.2 µmole 1 16 1440 µL
2 14 1440 µL
1.0 µmole 1 11 1392 µL
2 10 1440 µL