Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA, U.K.
Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge CB2 3EG, U.K.
ACS Synth Biol. 2020 Apr 17;9(4):864-882. doi: 10.1021/acssynbio.9b00511. Epub 2020 Apr 2.
We present the OpenPlant toolkit, a set of interlinked resources and techniques to develop Marchantia as testbed for bioengineering in plants. Marchantia is a liverwort, a simple plant with an open form of development that allows direct visualization of gene expression and dynamics of cellular growth in living tissues. We describe new techniques for simple and efficient axenic propagation and maintenance of Marchantia lines with no requirement for glasshouse facilities. Marchantia plants spontaneously produce clonal propagules within a few weeks of regeneration, and lines can be amplified million-fold in a single generation by induction of the sexual phase of growth, crossing, and harvesting of progeny spores. The plant has a simple morphology and genome with reduced gene redundancy, and the dominant phase of its life cycle is haploid, making genetic analysis easier. We have built robust Loop assembly vector systems for nuclear and chloroplast transformation and genome editing. These have provided the basis for building and testing a modular library of standardized DNA elements with highly desirable properties. We have screened transcriptomic data to identify a range of candidate genes, extracted putative promoter sequences, and tested them to identify new constitutive promoter elements. The resources have been combined into a toolkit for plant bioengineering that is accessible for laboratories without access to traditional facilities for plant biology research. The toolkit is being made available under the terms of the OpenMTA and will facilitate the establishment of common standards and the use of this simple plant as testbed for synthetic biology.
我们展示了 OpenPlant 工具包，这是一组相互关联的资源和技术，用于将 Marchantia 开发为植物生物工程的测试平台。Marchantia 是一种苔类植物，是一种具有开放发育形式的简单植物，允许在活体组织中直接可视化基因表达和细胞生长的动态。我们描述了用于简单高效的无菌繁殖和维持 Marchantia 系的新技术，这些技术不需要温室设施。Marchantia 植物在再生后几周内会自发产生克隆繁殖体，并且可以通过诱导生长的有性阶段、杂交和收获后代孢子，在单个世代中扩增百万倍。该植物具有简单的形态和基因组，基因冗余度降低，其生命周期的显性阶段是单倍体，这使得遗传分析更加容易。我们已经构建了用于核和叶绿体转化和基因组编辑的稳健 Loop 组装载体系统。这些为构建和测试具有高度理想特性的标准化 DNA 元件的模块化库提供了基础。我们已经筛选了转录组数据，以鉴定一系列候选基因，提取了推定的启动子序列，并对其进行了测试，以鉴定新的组成型启动子元件。这些资源已被整合到一个植物生物工程工具包中，可供没有传统植物生物学研究设施的实验室使用。该工具包将根据 OpenMTA 的条款提供，这将有助于建立共同的标准，并将这种简单的植物用作合成生物学的测试平台。
