Large DNA molecules are synthesized by the enzymatic assembly of short DNA oligonucleotide 60-100bp in length. Currently, each DNA fragment is synthesized in relatively small numbers at an excessive macroscopic scale, incurring a large manufacturing overhead in production costs. It is possible to make these building blocks cheaply by building them on microarrays using controlled light or electronic arrays. However, microarray DNA is both extremely error-prone and comes as dilute, hypercomplex mixtures of tens of thousands of different species. To effectively use a large microarray library, they need to be sorted apart from one another and filtered such that only correctly synthesized pieces are used for the final assembly steps. We have built a system that does this physically using next-generation sequencers that sequence copies on beads of clonal DNA species sampled from the microarray. We then use high-speed pulsed lasers to eject the correct beads of each sequence into unique, specified combinations in 384-well plates for assembly into genes. We propose to develop an enzymatic pipeline to convert our sequenced, ejected beads into kilobase-sized fragments. This requires developing reactions for amplification, processing, and assembly of our sequenced ejection material.
Keywords: DNA synthesis, Synthetic Biology, Synthetic Metagenomics, Sequencing, Microarray
