by Steven Burgess and Cindy Chan
How researchers answer questions around innovation, patenting and open access will have a profound impact on the development of the synthetic biology community. These issues were at the center of discussions during the OpenPlant Forum last month as researchers gathered at the John Innes Centre to present the latest progress in plant synthetic biology. Here we provide a review of the seven key developments presented at the meeting.
1. Intellectual Property, Open Science and Innovation
Patents are the main form of intellectual property (IP) and Oliver Sexton from the Rainbow Seed Fund emphasized the importance of IP when start-up companies seek to raise funding from venture capitalists.
— Cindy Chan (@cindyckx) July 25, 2016
However, there are alternatives, and there was some concern that patents are often used as blocking agents to stifle innovation. Dominic Berry of the Engineering Life team talked about what the synthetic biology community might be able to learn from the development of breeder’s rights, which includes an exemption that allows the use of protected varieties to develop novel strains.
Following on from this Jenny Molloy introduced the efforts of Linda Kahl and the OpenPlant IP Working Group to develop an Open Material Transfer Agreement (OMTA). MTAs are commonly used, non-patent forms of IP, but as indicated in the ‘Use and Misuse of MTAs’ current formats suffer from a number of disadvantages and are rarely ever enforced. An OMTA could allow for commercial reuse and the transfer of materials to third parties to promote collaboration and innovation. Currently discussions are ongoing between the University of Cambridge, Stanford and institutes at the Norwich Research Park to pilot its use, and if successful it may become more widely used.
Video: OxSyBio 3D printed network of cells folding up
A move to more open ways of starting and building companies is a growing trend. Tom Meany talked about how engaging with the DIYbio movement stimulated ideas for startup companies during the UK’s First Bio-Hackathon, with the winners entered onto the Judge Business School’s Accelerate Cambridge Program to develop software to facilitate automated DNA construct assembly, and startup company OxSyBio made a call for academic collaborators to identify new uses of their precision cell printing technology to grow their business.
— Transcriptic (@transcriptic) June 23, 2016
A critical mass of active and engaged participants is essential to effectively drive innovation using open technologies, and Fernan Federici talked about creating a community of synthetic biology developers in Latin America. He highlighted both the growth of the TECNOx competition which provides an alternative to iGEM, as well developing opensource technologies, giving the example of a low cost method to measure fluorescence developed with BackyardBrains that could be particularly useful for educational purposes.
2. Marchantia as a Model System for Plant SynBio
To address some of the issues raised above, Prof. Jim Haseloff outlined the two tiered approach to intellectual property adopted by the OpenPlant project, whereby the liverwort Marchantia polymorphia is being developed as system to provide open source tools to drive innovation in plant systems, while the potential to generate patents in crop species is retained.
Marchantia, revolutionizing plant synbio. Source: CC BY-SA 3.0, https://commons.wikimedia.org/w/index.php?curid=17208
To achieve this goal, there has been a push to establish Marchantia as a chassis for plant synthetic biology. Susana Sauret-Gueto gave an update on efforts to setup community resources for Marchantia, including flexible liquid handling and imaging pipelines that can be adapted for small scale small projects and high throughput experiments, as well as transformation techniques, short-term & long-term storage of transgenic lines, spore production and an enhancer trap screen.
— Jim Haseloff (@jimhaseloff) July 28, 2016
The breadth of Marchantia research was reflected in the presentations given. Bernado Pollack talked about developing automated spatial ratiometric analysis of Marchantia, Mihails Delmans presented MarpoDB.io for exploring Marchantia genomic data, Guru Radhakrishnan introduced Marchantia paleacea as a novel system for studying the evolution of Arbuscular mycorrhizal symbiosis and Orr Yarkoni presented his work on redesigning the Marchantia chloroplast genome, including synthesis of codon optimized fluorescent proteins.
3. Standards and Reproducibility
The need for better documentation and adherence to standards was a recurring theme throughout the two days. The idea of introducing short bioRxiv publications for DNA parts was introduced as a way of encouraging sharing, improving quality and recognizing outputs through provision of a referenceable DOI.
bioRxiv is an open access preprint server run by Cold Spring Harbor laboratory. http://biorxiv.org/
Additionally Tobias Wenzel talked about the DocuBricks platform for documenting open hardware projects in accordance with the Gathering For Open Science Hardware (GOSH) manifesto and a panel discussion highlighted efforts to establish an effective means for sharing protocols to help tackle the reproducibility problem in science. After reviewing a range of platforms Protocols.io was proposed as the best option for sharing procedures, the site allows version control, so it is possible to see how protocols evolve, “forking”, so people can copy and modifying existing procedures, comments to identify technical issues and provide a DOI to reference.
Richard Bowman’s Waterscope was highlighted as one of the best documented open hardware projects available on Docubricks. 12/05/16 Waterscope is a new company developing a cheap, lightweight system to test water quality in the developing world. Picture: Keith Heppell
4. Automation for SynBio
Modularity is key to automation, and the ability to share and re-use DNA parts will be necessary to accelerate DNA construct assembly. As a result, Nicola Patron previously led community efforts to adopt a common syntax for MoClo golden gate DNA assembly in plants, and at the meeting Francisco Navarro talked about expanding this system for use in the model green alga Chlamydomonas reinhardtii.
Chlamydomonas reinhardtii By Dartmouth Electron Microscope Facility, Dartmouth College – Source and public domain notice at http://remf.dartmouth.edu/imagesindex.html, Public Domain, https://commons.wikimedia.org/w/index.php?curid=2787116
Golden-gate DNA assembly allows for construction of large multi-gene constructs, however correct regulation of genes within these vectors is an area of active research. Looking to the future, Hans-Wilhelm Nutzmann emphasized the importance of the chromatin landscape in controlling Metabolic Gene Clusters in plants, and highlighted a better understanding of these processes may help improve vector design.
5. Data Mining For Synbio
Multi-gene constructs have been particularly useful in the field of metabolic engineering, where it is often necessary to introduce several novel enzymes to complete a synthetic pathway. There is commercial interest in utilizing biological organisms for the production of chemicals, and this is underpinned by an understanding of the enzymes available in nature. Peter Murray-Rust presented the ContentMine software which may aid this process by extracting information about phytochemicals from the plant literature. The usefulness of mining publicly available data was emphasized by Tom Knight when he went someway to explaining just what anyone does with 600Mbp of DNA in his talk about the synthetic biology company Ginkgo Bioworks.
Ginkgo use yeast for the production of fragrances and high value compounds, and in order to develop biosynthetic pathways for this, they have developed methods to mine databases and cluster sequences in as a means of predicting enzyme activity. Ginkgo then order libraries of enzyme sequences and use a combination of semi-intelligent modification of active sites and an in vitro screening platform to identify the best enzymes for each step in the pathway, before assembling them in vivo.
— OpenPlant (@_OpenPlant) July 27, 2016
6. Metabolic Engineering
Continuing on the theme of metabolic engineering there were a variety of talks about harnessing plants for chemical production. Allan Green of CSIRO and Johnathan Napier of Rothamsted Research presented projects to produce omega-3-fatty acids in plants using Canola and Camelina respectively. These chemicals are important for both aquaculture and human health, and are only synthesized by algae. As a result, production of omega-3-fatty acids in plants requires the addition of 7 genes from algae, and after almost 20 years of research both projects have reported yields in excess ~15% seed mass. This means for example that 1HA of canola can provide the equivalent amount of DHA as 10,000 fish. Both crops are currently in in field trials – Canola as part of collaboration between CSIRO, GRDC and NuSeed in Australia and Camelina in the UK.
Allan Green also presented CSIRO’s efforts to engineer safflower as a platform for production of high purity oleic acid, which can be used as a precursor molecule for the synthesis of biolubricants and nylon. As oils are typically only produced in seeds at significant levels much of the biomass is wasted and Green finished up by showing work at CSIRO to increase oil content in potato tubers and leaves of tobacco, the latter of which would cost $100-200/barrel with current yields.
The approach of utilizing of all parts of plant biomass for commercial means will be vital to any future efforts to reduce production costs. Progress in this direction was demonstrated by Patrick Jarvis of Associated British Foods plc (ABF) who presented the Wissington Sugar Biorefinery for processing sugar beet, which produces only 3 skips of waste in an entire year.
In addition to adding heterologous biosynthetic pathways, there is interest in utilizing the innate ability of plants to synthesize a remarkable range of molecules. To achieve this aim Cathie Martin highlighted the importance of altering transcription factor abundance to boost the levels of entire metabolic pathways. Her group has taken this approach to produce purple tomatoes enriched with anthocyanin and increased production of flavonols in tomato to extend their shelf life.
When boosting endogenous metabolic pathways, the tissue specificity of processes becomes important to consider both in terms of plant physiology and to work out how to optimize metabolite production. To address this, Lorenzo Caputi presented advances in imaging techniques to visualize the spatial distribution of metabolic production and Ivan Reyna-Llorens talked about the importance of identifying novel promoters to control spatial regulation of gene expression as part of efforts to enhance the photosynthetic capacity of rice.
7. Genome Editing
Professor Wendy Harwood and Professor Alison Smith both presented the applications and challenges of using CRISPR Cas9 system in plants. Wendy Harwood demonstrated successful CRISPR Cas9 knockouts in crops such as barley and Brassica oleracea, but noted that the efficiency of process requires improvement. Alison Smith, on the other hand, provided insight into the of production of ‘resistant starch‘ which can reduce calorie consumption, giving the example of successful production of starch synthase RNAi knockouts in Arabidopsis thaliana. Following this, they hope to create similar gene knockouts in potatoes using a CRISPR-Cas9 system, which could be used to create low calorie potato products.
CRISPR Crisps anyone? Image by Evan-Amos – Own work, Public Domain, https://commons.wikimedia.org/w/index.php?curid=11926930
For screening purposes, transient expression systems offer a valuable alternative to stable transformation of plants and Professor George Lomonossoff discussed the applications of hypertrans technology which will be used by Leaf Systems as part of their service for transient expression profiling in Nicotiana benthamiana. Hypertrans involves use of a modified Cow-Pea Mosiac Virus (CPMV)-HT protein expression system, Lomonossoff provided example applications such as producing plant-based poliovirus vaccines and understanding of the biosynthesis of antimicrobial triterpenoids (avenacins) in Oats.
Aymeric Leveau of the Obsorne Lab expanded on the later project and presented his work to transfer the biosynthetic pathway of Avenacins from Oats into wheat, in order to protect against Gaeumannomyces graminis var tritici infection that is responsible for the damaging “Take-all” disease.
Over the two days the forum provided a stimulating mix of science and business. However, as researchers are looking to engineer plants there were also many discussions about the importance of regulation and ‘public perception’. Former SynBioLEAP fellow Wieke Betten and lecturer in ethics, emphasized the fact that ‘the public’ is made up of many different interests groups with different perspectives, urging the importance of understanding the views of all stakeholders when taking any regulatory decision.
— Wieke Betten (@ouique) July 27, 2016
Additionally, Dominic Berry pointed out that it is a common misconception that concerns about GM crops are based on safety, when it in reality most worries resolve around who is going to benefit from new GM varieties and whether or not they are better than other approaches.
In summary, as well as showcasing excellent science, the forum highlighted the importance of many issues around intellectual property and regulation that will effect the wider synthetic biology community. It was encouraging to see researcher’s engaging with these broader issues, as was particularly evident in Rothamstead Research’s decision to be proactive in engaging with the public during their field trials of GM Camelina.
Discussions around opensource vs patenting, regulation vs developing technologies will continue, but what the forum demonstrated is that they don’t have to be mutually exclusive.
Disclaimer: Steven Burgess and Cindy Chan have both received funds from the OpenPlant project for research purposes.