CRISPR base editor-based targeted random mutagenesis (BE-TRM) toolbox for directed evolution.

IF 2.9 3区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

BMB Reports Pub Date : 2024-01-01

Rahul Mahadev Shelake, Dibyajyoti Pramanik, Jae-Yean Kim

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引用次数: 0

Abstract

Directed evolution (DE) of desired locus by targeted random mutagenesis (TRM) tools is a powerful approach for generating genetic variations with novel or improved functions, particularly in complex genomes. TRM-based DE involves developing a mutant library of targeted DNA sequences and screening the variants for the desired properties. However, DE methods have for a long time been confined to bacteria and yeasts. Lately, CRISPR/Cas and DNA deaminase-based tools that circumvent enduring barriers such as longer life cycle, small library sizes, and low mutation rates have been developed to facilitate DE in native genetic environments of multicellular organisms. Notably, deaminase-based base editing-TRM (BE-TRM) tools have greatly expanded the scope and efficiency of DE schemes by enabling base substitutions and randomization of targeted DNA sequences. BE-TRM tools provide a robust platform for the continuous molecular evolution of desired proteins, metabolic pathway engineering, creation of a mutant library of desired locus to evolve novel functions, and other applications, such as predicting mutants conferring antibiotic resistance. This review provides timely updates on the recent advances in BE-TRM tools for DE, their applications in biology, and future directions for further improvements. [BMB Reports 2024; 57(1): 30-39].

本刊更多论文

用于定向进化的基于 CRISPR 碱基编辑器的定向随机诱变（BE-TRM）工具箱。

通过定向随机诱变（TRM）工具对所需基因座进行定向进化（DE），是产生具有新功能或改进功能的遗传变异的有力方法，尤其是在复杂基因组中。基于 TRM 的定向进化涉及开发目标 DNA 序列的突变体文库，并筛选具有所需特性的变体。然而，长期以来，DE 方法一直局限于细菌和酵母。最近，人们开发了基于 CRISPR/Cas 和 DNA 去氨酶的工具，这些工具可以规避生命周期较长、文库规模较小和突变率较低等持久性障碍，从而促进多细胞生物原生遗传环境中的去氨化。值得注意的是，基于脱氨酶的碱基编辑-TRM（BE-TRM）工具通过对目标 DNA 序列进行碱基置换和随机化，极大地扩展了脱氧核糖核酸（DE）计划的范围和效率。BE-TRM工具为所需蛋白质的持续分子进化、代谢途径工程、创建所需基因座的突变体库以进化新功能以及其他应用（如预测赋予抗生素抗性的突变体）提供了一个强大的平台。本综述及时更新了用于 DE 的 BE-TRM 工具的最新进展、其在生物学中的应用以及未来进一步改进的方向。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

BMB Reports 生物-生化与分子生物学

CiteScore

5.10

自引率

7.90%

发文量

141

审稿时长

1 months

期刊介绍： The BMB Reports (BMB Rep, established in 1968) is published at the end of every month by Korean Society for Biochemistry and Molecular Biology. Copyright is reserved by the Society. The journal publishes short articles and mini reviews. We expect that the BMB Reports will deliver the new scientific findings and knowledge to our readers in fast and timely manner.