• Research

Automated Nanoscale DNA Assembly

We are working to push the limits of the size and scale of DNA assembly and have recently automated the fabrication of large DNA assemblies in sub-microliter volumes

Project summary.

Led by: Patron Group

Start date: June 2014

End date: September 2019

Duration: 61 months

Grants:

BBSRC Synthetic Biology Research Centre ‘OpenPlant’ Award (BB/L014130/1)

DNA Synthesis at Norwich Research Park (BB/M000966/1)

Funding:

Total award £13,690,968

The field of bioengineering is underpinned by our ability to assembly bespoke and complex DNA molecules at scale.  We are working to push the limits of the size and scale of DNA assembly and have recently automated the fabrication of large DNA assemblies in sub-microliter volumes at the Earlham DNA Foundry. This has been coupled to validation using next-generation sequencing technologies.

Details.

The primary costs of assembling DNA, especially as we move towards an era of synthetic genomics, are those associated with the purchase of synthetic DNA molecules, and those associated with constructing and sequence-verifying the assemblies.
 
The cost of assembling DNA fragments can be reduced by increasing throughput and reducing the input to the reaction. This can be achieved by miniaturizing and automating the reactions. Facilities, known as DNA Foundries, are dedicated to automating the design, construction and characterization of fabricated DNA.
 
We are using laboratory automation, including acoustic dispensing technologies to transfer nanoliter-scale liquid droplets with high precision and accuracy. This has enabled DNA assembly reactions to be set-up at the nanoliter scale, cutting reagent cost by up to 100-fold. This is coupled with robotic platforms for colony picking and purification of plasmid DNA, reducing the effort of the entire cloning workflow. Further, the cost of assembly verification and rigorous quality control has been implemented by replacing Sanger sequencing with automated, highly multiplexed next-generation sequencing of assembled constructs.

Publications.

DNA assembly standards: Setting the low-level programming code for plant biotechnology.

Vazquez-Vilar M, Orzaez D, Patron N, In press, Plant Science (2018)
DOI: 10.1016/j.plantsci.2018.02.0243

Loop Assembly: a simple and open system for recursive fabrication of DNA circuits

Pollak B, Cerda A, Delmans M, Álamos S, Moyano T, West A, Gutiérrez RA, Patron NJ, Federici F*, Haseloff J*, bioRxiv preprint (2018)
DOI: 10.1101/247593

SMRT Gate: A method for validation of synthetic constructs on Pacific Biosciences sequencing platforms

D’Amore R, Johnson J, Haldenby S, Hall N, Hughes M, Joynson R, Kenny JG Patron NJ, Hertz-Fowler C and Hall A*, Biotechniques 63 (2017)
DOI: 10.2144/000114565

Blueprints for green biotech: development and application of standards for plant synthetic biology

Patron, N.J., Biochemical Society Transactions 44, 702-708 (2016)
DOI: 10.1042/BST20160044

Collaborators.

The OpenPlant Consortium

OpenPlant is a collaborative initiative between the University of Cambridge, the John Innes Centre and the Sainsbury Laboratory in Norwich. The initiative promotes interdisciplinary exchange, open technologies for innovation and responsible innovation for sustainable agriculture and conservation. The Development and application of standards for plant synthetic biology was a collaborative aim of the consortium to underpin the efforts in all work-packages.

The Liverpool GeneMill

GeneMill specializes in the fabrication of synthetic DNA providing end-to-end support from the design stage through to building sequence verified constructs.

Impact statement.

Synthetic biology applies engineering principles such as standardisation and modularisation to biological sciences through application of iterative design-build-test-learn cycles. The standardisation of biological components and reactions allows workflows to be automated. This provides the potential to revolutionise the speed and scale of research, increasing accuracy and allowing miniaturisation to reduce costs. The adoption of these principles and approaches enables scientists to pursue complex experimental designs.

Related reading.