Progress in gene assembly from a MAS-driven DNA microarray

Abstract

We report the progress on 765 bp full green fluorescent protein (GFP) gene assembly of oligonucleotides from a single DNA microarray, using the amplification and assembly of chip-eluted DNA (AACED) strategy. The chip was fabricated with maskless array synthesizer (MAS) via 2-nitrophenylpropoxycarbonyl (NPPOC)-driven light-directed synthesis. Upon the recognition of the strong dependence of final assembly yield on the purity of construct oligonucleotides, we put a large effort in developing error reduction methods with a considerable improvement in the purity of chip-oligonucleotides in terms of exposure image quality, synthesis protocols and mask design. In addition to the synthesis improvement (as an error prevention), a post-assembly error filtering method was devised and developed – consensus shuffling using MutS-protein filtering. The preliminary demonstrations indicate their potential uses in more rapid and efficient DNA assembly process.