Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Groningen, The Netherlands.
Chembiochem. 2022 Apr 5;23(7):e202100507. doi: 10.1002/cbic.202100507. Epub 2021 Dec 7.
The construction of custom libraries is critical for rational protein engineering and directed evolution. Array-synthesized oligo pools of thousands of user-defined sequences (up to ∼350 bases in length) have emerged as a low-cost commercially available source of DNA. These pools cost ≤10 % (depending on error rate and length) of other commercial sources of custom DNA, and this significant cost difference can determine whether an enzyme engineering project can be realized on a given research budget. However, while being cheap, oligo pools do suffer from a low concentration of individual oligos and relatively high error rates. Several powerful techniques that specifically make use of oligo pools have been developed and proven valuable or even essential for next-generation protein and pathway engineering strategies, such as sequence-function mapping, enzyme minimization, or de-novo design. Here we consolidate the knowledge on these techniques and their applications to facilitate the use of oligo pools within the protein engineering community.
定制文库的构建对于理性的蛋白质工程和定向进化至关重要。由数千个用户定义序列组成的阵列合成寡核苷酸池（长度最长可达约 350 个碱基）已经成为一种低成本的、可商业获得的 DNA 来源。这些寡核苷酸池的价格比其他定制 DNA 的商业来源低≤10%（取决于错误率和长度），而这种显著的成本差异可能决定了一个酶工程项目是否可以在给定的研究预算下实现。然而，尽管价格低廉，寡核苷酸池确实存在单个寡核苷酸浓度低和相对较高错误率的问题。已经开发出了几种专门利用寡核苷酸池的强大技术，这些技术已经被证明对于下一代蛋白质和途径工程策略具有价值，甚至是必不可少的，例如序列-功能映射、酶最小化或从头设计。在这里，我们整合了这些技术及其应用的知识，以促进寡核苷酸池在蛋白质工程界的应用。
