10-nmol oligonucleotide synthesis for the ABI model 394 DNA synthesizer

Abstract

Sequencing and gene contig assembly generally involve primer walking, in which an oligonucleotide primer is used once and then discarded. Because the smallest commonly available scale for commercial oligonucleotide synthesizers is the 40-nmol scale, a large excess of product is produced. This results in reagent waste, excess cost to the researcher, and increased production time. A logical solution would be to develop a smaller scale for the commercial synthesizers. A 10-nmol scale was developed for the Applied Biosystems ABI model 394 DNA synthesizer. To develop the 10-nmol scale, a column support was first developed from commonly available materials. Then a 10-nmol cycle was developed from the systematic reduction of the reagents. The final product of the 10-nmol cycle was tested for quality by monitoring the coupling efficiency, polyacrylamide gel electrophoresis, and automated DNA sequencing.The final cycle reduces the cost of the oligonucleotide and decreases the time required to produce it.