摘要/Abstract
作为临床药物的重要来源, 放线菌天然产物及其衍生物已被大规模开发并应用于临床. 为了实现放线菌来源活性天然产物的高效合成、新型天然产物的结构衍生以及未知天然产物的定向挖掘, 往往需要聚焦于生物合成基因簇, 运用生物学技术开展理性的遗传改造. 规律性成簇间隔的短回文重复序列及相关蛋白(CRISPR/Cas)是部分细菌和古细菌在长期自然进化过程中发展而来的一种对抗外源遗传物质入侵的适应性免疫防御系统. 以CRISPR/Cas9为代表的基因编辑系统已被广泛应用于脱氧核糖核酸(DNA)片段的靶向特异性切割, 为生物技术领域的发展提供了无限可能. 综述了CRISPR/Cas9系统在放线菌遗传改造中的应用情况, 包括在基因簇抓取、编辑与重构、原位同源重组以及代谢调控方面的研究进展, 涉及全新的CRISPR/Cas9介导的基因编辑方法以及代表性的成功案例, 为高效利用CRISPR/Cas9系统获取放线菌天然产物提供了借鉴.
关键词: 放线菌, 天然产物, 生物合成基因簇, 基因编辑, 规律性间成簇间隔的短回文重复序列及相关蛋白9 (CRISPR/Cas9)
As important source of clinical drugs, the natural products in actinomycetes and their derivatives have been developed on a large scale and used in clinical practice. In order to achieve efficient synthesis of active natural products, structural derivation of new natural products, and targeted mining of unknown natural products, researches mainly focus on the biosynthetic gene clusters from actinomycetes, which should be rationally engineered by advanced biotechnology. CRISPR/ Cas, which represents the clustered regularly interspaced short palindromic repeats and the associated protein, is an adaptive immune system developed by some bacteria and archaea in the long-term natural evolution process to resist the invasion of foreign genetic materials. As a representative of gene editing system, the clustered regularly interspaced short palindromic repeats and the associated protein 9 (CRISPR/Cas9) has been widely used in the targeted editing of deoxyribonucleic acid (DNA) fragments, which provides unlimited possibilities for the development of biotechnology. This paper reviews the application of the CRISPR/Cas9 system in the genetic manipulation of actinomycetes, including the recent research progress in capture, editing and reconstruction of gene clusters, in situ homologous recombination and metabolic regulation, which involves the novel CRISPR/Cas9-mediated gene editing methods and the representative successful cases. This paper will provide valuable information for the efficient use of CRISPR/Cas9 to obtain natural products in actinomycetes.
Key words: actinomycetes, natural products, biosynthetic gene clusters, gene editing, the clustered regularly interspaced short palindromic repeats and the associated protein 9 (CRISPR/Cas9)
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