关键词: 细胞工厂快速构建, 自动化合成生物技术, DNA组装, 底盘细胞操作

Abstract:

The construction of microbial cell factories plays a critical role in green biomanufacturing. The production of chemicals can be achieved using microbial cell factories, instead of traditional production methods that rely on plants and land resources. In recent years, synthetic biology has made significant advancements in biological manufacturing, including the successful biosynthesis and commercialization of natural products derived from plants, such as artemisinin and cannabidiol. However, the construction of engineered microbial cell factories still faces challenges due to the high complexity of living systems, the iterative nature of experimental processes, and low experimental throughput. To overcome these obstacles, the application of high-throughput, automated, and intelligent hardware, and software platforms, as well as standard, modular libraries of synthetic biology parts and processes, has led to the emergence of automated synthetic biotechnology. This breakthrough allows for low-cost, high-throughput, rapid, and iterative experimentation to efficiently construct large-scale engineering prototypes of microbial cell factories, thereby facilitating related research and applications in this field. To this end, dozens of automated biofoundries have been built around the world. This article primarily focuses on the application of automated synthetic biotechnology in the most critical and time-consuming step of "build" in the "design-build-test-learn" cycle of synthetic biology. The automated building of microbial cell factory demands both the automated construction of engineered DNA and the automated manipulation of microbial chassis, including gene synthesis, PCR amplification, enzymatic digestion, DNA assembly, transformation and plating, colony picking, cell lysis, nucleotide purification, and sequencing. In this review, we summarized the state-of-the-art automated processes and platforms for DNA assembly and microbial chassis manipulation, provided recent advances of automated synthetic biotechnology in the mining and characterization of biosynthetic gene clusters, the combinatorial optimization of metabolic pathways, and the engineering of microbial chassis. We also discussed the challenges and opportunities in automating the construction of microbial cell factories, putting forward to one of most important future directions in full process automation, including the construction of non-model microbial cell factories. The localization and independent research and development of automated biofoundry would make significant contributions to this process.

Key words: rapid construction of cell factories, automated synthetic biotechnology, DNA assembly, chassis manipulation