Working with the liverwort Marchantia polymorpha, we are the first team with the privilege of being able to generate stable plant transformants, grown with little effort on agar plates.

As such, throughout our iGEM experience, we had two main goals. The first was to develop a project that had a real and valuable application and which would work ''with'' the complex multicellular machinery that nature hands down to us in Marchantia. Our second, and perhaps more important, goal, was to build a foundation for future engineering of Marchantia, both for iGEM teams and the wider scientific community.

mösbi, the Marchantia Open Source Biosensor, was a project that enabled us to bring these two together. The idea is simple: mösbi is made up of input, processing and output modules. These modules communicate by way of the transcription factors GAL4 and HAP1. mösbi is truly open source: modules can be developed completely independently of each other, so long as they adhere to the simple framework. They can be 'installed' either in the lab, by direct Agrobacterium Mediated Transformation, or at home, by growing up plants with individual modules built in, and then simply crossing them. A biosensor (which detects, for example, heavy metals in water) which can be shipped as spores, grown and propagated with virtually no resources, speaks for itself in terms of value to those without need for expensive equipment to assay their essentials. Furthermore, the innovative use of Marchantia's inbuilt haploid reproduction machinery - that is, Mendelian crossing, provides a unique meeting point for synthetic biology in the lab and at home. We think of mösbi as one of the first instances of consumer biotechnology.

Having established our input-processing-output framework, we structured our lab work to develop each plugin type in parallel. We sought to assay in Marchantia each module as standalone consruct - constitutively active, or with simple fluorescent reporters, and as components of a mösbi circuit.

As outputs, we considered chromoproteins, which would make both extremely easy sensing readouts and useful reporters for plant synthetic biology research, as well as the synthesis of raspberry ketone from p-coumaroyl coenzyme A, which would be proof of concept that more complicated metabolic circuits can be integrated as outputs for mösbi. We gained evidence that some of the chromoproteins, which we received from the Uppsala team 2013, were succesfully transformed, but not expressed, and that asPink was expressed, but not visible to the naked eye in the background of marchantia's vibrant colours. We quantitatively characterised a three enzyme raspberry ketone pathway, components of which we received from Ikuro Abe, Jules Beekwilder and Esmer Jongedijk, in an odorless strain of E. coli from D. Summers lab, and transformed Marchantia with our two enzyme construct, but from our quantitative work in E. coli, we did not expect to observe any noticeable odors. We built a limonene synthesising construct, which we unfortunately did not receive the opportunity to transform into Marchantia. We designed an enhancer trap, which will enable us to identify Marchantia lines with expression patterns that would enable multiple outputs from different regions of the plant. We transformed Marchantia with the enhancer trap construct, and are waiting for them to reach a stage where they can be screened. These lines will be able to have mösbi output modules directly crossed in, and will be of great value to the developmental study of Marchantia.

Needing inputs for mösbi and a collection of promoters for our new chassis, we performed an extensive phylogenetic hunt for endogenous promoters from Marchantia genome/transcriptome data. We isolated these from the Marchantia genome, and assembled them into standalone and mösbi-compatible constructs. During our bioinformatic analysis, we produced for the first time a Marchantia codon table for optimizing expression. We also built a heat-shock promoter mösbi-module, from the DNA provided by Christian Boehm, which would enable smart environmental sensing in Marchantia. We were fascinated by hammerhead ribozymes, which could be evolved in and isolated from S. cerevisiae to control cleavage of RNA transcript, and hence expression, in response to a given ligand - and which could be cascaded to create logic elements. These hadn't yet been used in plants. We kindly received samples from the Smolke lab, and assembled these into standalone constructs and mösbi-processing/input modules.Unfortunately, problems in the later stages of transformation prevented us from assaying our constructs in vivo, and incompatibility with RFC10 prevent us from submitting these to the registry. We submitted many of the candidate promoters, along with new reporter localisation sequences and fundamental Marchantia tools as digital parts to the registry.

We struggled at various stages with assembling our constructs. They usually were large, and consisted of many parts, and came from a range of sources. To try to improve our efficiency on larger constructs, we successfully tested the newly optimized Ligase Cycling Reaction, and used it to build some of our larger constructs. However, this came at a great time cost. We realised that to truly make use of the rapid transformation cycle of Marchantia, and to enable future members of the plant synthetic biology community, a method of sharing reusable plant parts was essential. To pave the way for this, we collaborated with Valencia and UEA-NRP, the only other iGEM teams working in plants, to author BBF RFC 105, which sets up a standardised PlantSyntax for plant parts. We submitted mösbi-modules to the registry that enable the assembly techniques described in the paper, to insert input promoters, and reporters into module cassettes.

Appreciating that consumer biotechnology/synthetic biology in the home currently has many questions to answer on issues of safety and practice, we realised that it was crucial for us to assess these issues in the context of mösbi. We researched these issues and produced a paper addressing them as a way of demonstrating our responsibility. Acknowledging our responsibility to address these issues beyond the scientific community, we will also be hosting a session in the annual Cambridge Science Festival on October 26th, at the Cambridge Science Centre. We aim to educate the wider public about the potential of plant synthetic biology and Marchantia, as well as to address their concerns on genetic modification. We hosted students from the St. Paul's School collegiate iGEM team to inform them about practical aspects of iGEM, mösbi, and wider issues in the synthetic biology community.

We realised that we have the opportunity to truly impact the future access to Marchantia and plant synthetic biology more broadly, and that future uptake depends greatly on the quality of the resources we provide. As a result, we built the only non-standard laboratory equipment for Marchantia: a portable growth facility, and provided detailed instructions for future users to build their own. We will also lead a plants workshop, in collaboration with the Norwich-NRP and Valencia teams, at the Giant Jamboree. Collecting the expertise we gained from the Haseloff lab, we produced a comprehensive guide on Marchantia synthetic biology, which we hope will be openly added to by new users and the scientific community.

We hope that you agree that developing mösbi, the first sensor that can be grown at home and the first piece of consumer biotechnology, and truly working with all levels of the community to build a foundation for Marchantia synthetic biology are contributions in the true iGEM spirit.

We finish by expressing our sincerest thanks to our sponsors, those who donated their lab's work for us to work with, our wider support community at the Haseloff lab, the University of Cambridge and the John Innes Center. Finally, we give our personal thanks to Jim Haseloff, Bernardo Pollak, Anton Kan and Gos Micklem, whose long hours of effort with the team made for a truly wonderful iGEM experience.