An open-source advanced maskless synthesizer for light-directed chemical synthesis of large nucleic acid libraries and microarrays

Large- to ultra-large-scale synthesis of nucleic acids is becoming an increasingly important tool for understanding and manipulating biological systems, as well as for developing new technologies based on engineered biological materials, including DNA-based nanofabrication, aptamers and writing digital data at the molecular level. Several technologies for large-scale synthesis have been developed over the years, but all of them remain inaccessible to most researchers due to their complexity and/or use of proprietary technologies. Here, we present a fully open source, benchtop device for ultra-large scale nucleic acid synthesis that is also highly flexible and adaptable, able to accommodate a wide range of monomers and chemistries, while providing unrestricted access to synthesis parameter space

High-Efficiency Reverse (5′→3′) Synthesis of Complex DNA Microarrays

DNA microarrays are important analytical tools in genetics and have recently found multiple new biotechnological roles in applications requiring free 3′ terminal hydroxyl groups, particularly as a starting point for enzymatic extension via DNA or RNA polymerases. Here we demonstrate the highly efficient reverse synthesis of complex DNA arrays using a photolithographic approach. The method is analogous to conventional solid phase synthesis but makes use of phosphoramidites with the benzoyl-2-(2-nitrophenyl)-propoxycarbonyl (BzNPPOC) photolabile protecting group on the 3′-hydroxyl group. The use of BzNPPOC, with more than twice the photolytic efficiency of the 2-(2-nitrophenyl)-propoxycarbonyl (NPPOC) previously used for 5′→3′ synthesis, combined with additional optimizations to the coupling and oxidation reactions results in an approximately 3-fold improvement in the reverse synthesis efficiency of complex arrays of DNA oligonucleotides. The coupling efficiencies of the reverse phosphoramidites are as good as those of regular phosphoramidites, resulting in comparable yields. Microarrays of DNA surface tethered on the 5′ end and with free 3′ hydroxyl termini can be synthesized quickly and with similarly high stepwise coupling efficiency as microarrays using conventional 3′→5′ synthesis.© The Author(s) 201