Abstract:

At the beginning of this century, high-throughput in-parallel fermentation (cell culture) technology and its related apparatus based on microbioreactor and miniature bioreactor were developed and widely applied. This was to solve the challenges encountered by biopharmaceutical process researches in terms of cultivation throughput, R&D efficiency, and the continuous rising on the cost for microbial fermentation and mammalian cell cultures of multiple clones, and more importantly, the urgent needs for conducting design of experiment (DoE) driven by the principle of quality by design (QbD). In the recent years, the rapid advance of microbial metabolic engineering and synthetic biology has put forward a rather strong demands for the high-throughput screening on high-performance strain libraries and the early evaluation of strain phenotypic potential performance under industrial cultivation conditions. This has further expanded the application range of in-parallel fermentation technology with various culture volumes in the field of modern industrial biotechnology. Up to now, integrated systems containing in-parallel fermentation setups with multiple microbioreactors, online probes, operating software, and data processing units, which can simulate industrial cultivation conditions and implement accurate control of process parameters, have become a powerful tool for accelerating bioprocesses engineering R&D. They play a central role in transforming basic research achievements, such as drug discovery, metabolic engineering, and synthetic biology, into industrial technologies. Especially in the field of synthetic biology, based on the principle of "industrial similarity", in-parallel fermentation technology addresses key limitations of the conventional microplate and shake flask based high throughput screening, which cannot characterize the significant impact of culture conditions on the phenotypic performance of the selected clones at a large scale. This enables a process-oriented, high throughput, and efficient screening and evaluation if microbial strain libraries. This review provides an overview on the recent development in high throughput in-parallel fermentation & cell culture technology and its application scenarios of synthetic biology researches. In particular, it emphasizes the value of in-parallel fermentation technology in high-throughput strain screening complying with the three-stage strategy, and how in-parallel fermentation technology makes the implementation of the industrial similarity principle of strain screening possible, and how the technology combines DoE experimental tactic to significantly improve the efficiency of bioprocess development. It is also discussed the general procedure to build up a multivariate batch model of bioprocess based on in-parallel fermentation technology. In the end the approach of using in-parallel culture to establish a process scale down model is also explored.

Key words: fermentation, cell culture, in-parallel cultivation, high throughput, synthetic biology, design of experiment (DoE)

摘要：

21世纪初，为解决生物医药过程工程研究所面临的微生物和哺乳动物细胞培养的实验通量、研发效率与成本方面的问题，更重要的是质量源于设计（QbD）导向的生物过程工程实验设计（DoE）的迫切需要，基于微、小型生物反应器的平行发酵（细胞培养）技术与产品得到了广泛应用。近年来微生物代谢工程与合成生物学的飞速发展，对高性能菌种库的高通量筛选与菌种表型过程表现的早期评价提出了更高实验通量的需求，这进一步拓展了不同培养体积的平行发酵培养装置在工业生物技术领域的应用。时至今日，可模拟工业培养条件并实施过程参数准确控制的微小型反应器的多联罐平行发酵装置、系统操作软件和数据处理的集成系统已成为生物过程工程研发的强大工具，它在生物医药创新、代谢工程和合成生物学等基础研究成果向工业化技术转化中起到重要的支撑作用。特别是在合成生物学领域中，基于“工业相似性“原则的平行发酵技术，可以解决培养板或摇瓶高通量菌种筛选无法表征克隆表型、在规模化培养中的表现受培养过程参数显著影响的痛点问题，实现过程工程导向的高通量、高效率的菌种筛选与评价。本文对高通量平行发酵与细胞培养技术的发展近况与其在合成生物学研究中的应用场景做了介绍，其中主要总结了平行发酵培养技术在高通量菌种筛选评价“三段论”中的价值、平行发酵培养如何支持菌种筛选的工业相似性原则的实施、平行发酵培养结合DoE实验策略实施高效的生物过程工程开发、使用平行发酵培养建立过程多变元批次模型的方法，以及平行发酵培养与建立生物培养过程缩小模型的关系等。

关键词: 发酵, 细胞培养, 平行培养, 高通量, 合成生物学, 实验设计（DoE）