An automated linker-based DNA assembly standard

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Arturo Casini, James MacDonald, Tom Ellis, Geoff Baldwin

Imperial College London, United Kingdom

Cloning, and assembling DNA in general, is one of the main bottlenecks slowing down the progress of synthetic biology: it’s a slow and expensive process, and requires a lot of “ad hoc” adjustments. The lack of a standard also complicates exchange and comparison of constructs between different labs, as the same functional parts can be assembled in radically different ways. We propose a DNA assembly standard compatible with both the “small parts to genes” and the “genes to pathways” level. In order to achieve high throughput and improve reliability, this method is designed to be compatible with automated liquid-handling platforms and uses extremely robust reactions such as restriction digestions and ligations. It is also compatible with a wide range of part sizes and features (such as high GC% or presence of secondary structures) and can be used for the combinatorial assembly of libraries of parts. One of the main defining features of our approach is the use of “linker” DNA sequences to join parts together: they are the ones which participate in the assembly reactions and are computationally designed to achieve maximum efficiency and specificity.