Handbook of Single-Cell Technologies
Handbook of Single-Cell Technologies
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单细胞技术手册
DOI：
10.1007/978-981-10-8953-4_47
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发表时间：
2022
期刊：
影响因子：
--
通讯作者：
Gielen F
中科院分区：
文献类型：
--
作者：
Gielen F
酶负责维持生命的许多化学反应，并用于许多行业。实验室进化实验可以模拟达尔文的突变和选择压力过程，产生新进化的催化剂。最近开发的单细胞微胶囊技术能够筛选超大型酶突变体库，大大增加了分离改良催化剂的几率。封装过程使用液滴微流体技术，这是一种突破性的生化分析小型化和自动化技术，已经证明了其在进化酶和识别环境中的超微量催化剂方面的有效性。本章回顾了如何大规模的单细胞检测提供了一个有效的途径，对新的或改进功能的生物催化剂的鉴定。单细胞微囊化能够将基因型（基因序列）和表型（酶功能）保留在稳定的微区室中。在实践中，由单细胞表达的单基因突变体与其相应的酶被封装到单分散油包水微滴中，在其中进行测定。这使得在一天内筛选超大的108个成员的文库，与常规工具相比，同时降低了1000倍的测定试剂成本和增加了1000倍的通量。该技术已被广泛采用，用于加速发现和发展用于许多应用领域的生物催化剂，从污染环境的净化，生物能源生产，到塑料的降解和抗菌剂的发现。
Enzymes are responsible for many chemical reactions which support life and are used in numerous industries. Laboratory evolution experiments can mimic the Darwinian process of mutation and selection pressure, yielding newly evolved catalysts. Recently developed single-cell microencapsulation techniques enable the screening of ultralarge enzyme-mutant pools, greatly increasing the odds of isolating improved catalysts. The encapsulation process uses droplet microfluidics, a groundbreaking technology for the miniaturization and automation of biochemical assays which has already demonstrated its effectiveness at evolving enzymes and identifying ultrarare catalysts from the environment. This chapter reviews how large-scale single-cell assays provide an efficient route toward the identification of biocatalysts with novel or improved function. Single-cell microencapsulation enables retention of genotype (gene sequence) and phenotype (enzyme function) within a stable microcompartment. In practice, single-gene mutants expressed by single cells with their corresponding enzyme are encapsulated into monodisperse water-in-oil microdroplets where the assay takes place. This enables the screening of ultralarge 108members libraries in a day with a concomitant reduction of 1000-fold in assay-reagent costs and a 1000-fold increase in throughput compared to conventional tools. This technology has already been widely adopted for the accelerated discovery and evolution of biocatalysts used in many application domains from the decontamination of polluted environments, bioenergy production, to the degradation of plastics and the discovery of antimicrobials.