Automated DNA Assembly
As part of a collaboration between the University of Cambridge, Earlham Institute and the Universidad Católica de Chile, Pollak and Federici have devised a new method for gene assembly based on two Type IIS restriction endonuceases, BsaI and SapI. Loop Assembly allows rapid and efficient production of large DNA constructs, is compatible with widely used Level zero (L0) DNA parts such as Phytobricks, and can be easily automated.
Loop Assembly requires the alternating use of two Type IIS enzymes, Bsa1 (6-base-pair recognition sequence, 4 base overhang) and Sap1 (7 base-pair recognition sequence, 3 base overhang), and two sets of complementary plasmid vectors that allow efficient and ordered construction of 1, 4, 16, 64 gene fragments.
Roboticised construction of plant genes
Like other "Golden-gate"-based protocols, Loop Assembly does not require purification of individual DNA fragments, side products are recut during the ligation reaction to drive efficient formation of end-products. Loop Assembly is well suited to automation. OpenPlant researchers at the Earlham Institute and Cambridge are developing methods using acoustic-focusing non-contact liquid handling robots, which increases speed and scale of assembly, while reducing consumable costs and allowing reactions to be performed in nanolitre volumes.