Jones Sally, Martella Andrea, Cai Yizhi
Manchester Institute of Biotechnology (MIB), School of Chemistry, The University of Manchester, Manchester, United Kingdom.
Discovery Biology, Discovery Sciences, IMED Biotech Unit, AstraZeneca, Cambridge, United Kingdom.
Methods Enzymol. 2019;617:463-493. doi: 10.1016/bs.mie.2018.12.017. Epub 2019 Feb 13.
Construction of expression vectors is imperative for many areas of biological research and the biotechnology industry. Modular cloning systems for expression vector construction offer a labor- and cost-effective alternative to overcome drawbacks associated with traditional cloning methods. We developed an Extensible Mammalian Modular Assembly toolkit (EMMA) as an efficient and versatile tool to facilitate the construction of functionally diverse mammalian expression vectors from a standardized library of DNA parts. This system supports both hierarchical and combinatorial assembly, and is adaptable for automation. In this chapter, we describe the protocols to construct libraries of DNA parts and assemble these parts into mammalian expression vectors using EMMA.
构建表达载体对于生物学研究的许多领域和生物技术产业来说至关重要。用于构建表达载体的模块化克隆系统提供了一种经济高效的替代方案,以克服传统克隆方法的缺点。我们开发了一种可扩展的哺乳动物模块化组装工具包(EMMA),作为一种高效且通用的工具,用于从标准化的DNA元件库构建功能多样的哺乳动物表达载体。该系统支持分层组装和组合组装,并且适用于自动化操作。在本章中,我们描述了构建DNA元件库以及使用EMMA将这些元件组装成哺乳动物表达载体的方案。