Ribeiro Ana Luísa, Mencía Mario, Hidalgo Aurelio
Department of Molecular Biology, Center for Molecular Biology "Severo Ochoa" (UAM-CSIC), Universidad Autónoma de Madrid, Nicolas Cabrera 1, Madrid, 28049, Spain.
Methods Mol Biol. 2018;1685:131-143. doi: 10.1007/978-1-4939-7366-8_7.
The process of protein production optimization requires time and labor, constituting one of the main bottlenecks for the downstream utilization of the proteins. However, once through this bottleneck, the protein production process can be easily standardized and multiplexed to find the fittest variants in large libraries created by random mutagenesis. In this chapter, we present an overview of the most important choices to achieve homogeneous and functional expression of directed evolution libraries in microplate format: (1) choice of induction system and host strain, (2) choice of media and growth conditions, and (3) modifications to the genetic sequence.
蛋白质生产优化过程需要时间和人力，这构成了蛋白质下游利用的主要瓶颈之一。然而，一旦突破这一瓶颈，蛋白质生产过程就可以很容易地标准化和多路复用，以便在通过随机诱变创建的大型文库中找到最合适的变体。在本章中，我们概述了以微孔板形式实现定向进化文库均匀且功能性表达的最重要选择：（1）诱导系统和宿主菌株的选择，（2）培养基和生长条件的选择，以及（3）基因序列的修饰。
