Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, 9747 AG Groningen, The Netherlands.
Chair of Applied and Molecular Microbiology, Institute of Biotechnology, Technische Universität Berlin, 10623 Berlin, Germany.
ACS Synth Biol. 2021 Nov 19;10(11):2850-2861. doi: 10.1021/acssynbio.1c00260. Epub 2021 Nov 2.
Filamentous fungi are highly productive cell factories, often used in industry for the production of enzymes and small bioactive compounds. Recent years have seen an increasing number of synthetic-biology-based applications in fungi, emphasizing the need for a synthetic biology toolkit for these organisms. Here we present a collection of 96 genetic parts, characterized in or species, that are compatible and interchangeable with the Modular Cloning system. The toolkit contains natural and synthetic promoters (constitutive and inducible), terminators, fluorescent reporters, and selection markers. Furthermore, there are regulatory and DNA-binding domains of transcriptional regulators and components for implementing different CRISPR-based technologies. Genetic parts can be assembled into complex multipartite assemblies and delivered through genomic integration or expressed from an AMA1-sequence-based, fungal-replicating shuttle vector. With this toolkit, synthetic transcription units with established promoters, fusion proteins, or synthetic transcriptional regulation devices can be more rapidly assembled in a standardized and modular manner for novel fungal cell factories.
丝状真菌是高效的细胞工厂，常用于工业生产酶和小分子生物活性化合物。近年来，基于合成生物学的应用在真菌中越来越多，这强调了这些生物体需要合成生物学工具包。在这里，我们展示了一个包含 96 个遗传元件的集合，这些元件在 或 种中得到了表征，与模块化克隆系统兼容且可互换。该工具包包含天然和合成启动子（组成型和诱导型）、终止子、荧光报告基因和选择标记。此外，还有转录调控因子的调控和 DNA 结合结构域，以及用于实现不同 CRISPR 技术的组件。遗传元件可以组装成复杂的多部分组装体，并通过基因组整合或从基于 AMA1 序列的真菌复制穿梭载体进行表达。有了这个工具包，具有已建立启动子、融合蛋白或合成转录调控装置的合成转录单元可以以标准化和模块化的方式更快速地组装，用于新型真菌细胞工厂。
