摘要
丝状真菌(filamentous fungi)是广泛存在于自然界中的一类多细胞真核微生物,是酶制剂、有机酸、抗生素的核心生产体系,在生物技术中发挥着非常重要的作用。由于丝状真菌的生长发育较为复杂,其遗传体系和基因组编辑技术发展相对较慢,妨碍了丝状真菌基础研究和开发利用的快速发展。近年来,核酸酶介导的基因组编辑技术已经发展成为一种功能强大的基因组编辑工具,在生物技术领域的应用得到广泛关注,其中RNA介导的CRISPR系统(clustered regularly interspaced short palindromic repeats)已经成为新一代基因组编辑技术体系。文章将对目前使用最为广泛的3种基因编辑系统,包括CRISPR-Cas技术的发展历程进行概述,对CRISPR-Cas技术在丝状真菌基因组编辑中的研发进行系统介绍,包括在工业丝状真菌中的研发应用,例如鉴定次级代谢产物的关键基因并提高次级代谢物的生产能力、将外源基因定点整合到基因组上从而提高异源蛋白的表达、遗传重构蛋白分泌途径提高工业酶的产量等,最后对CRISPR-Cas基因组编辑技术及其衍生系统在丝状真菌的真菌基因功能研究、代谢途径重构、精确表达调控、蛋白定向进化以及高性能底盘构建等方面进行了展望。
Filamentous fungi,which are a large and diverse group of multicellular eukaryotic microorganisms existing in nature universally,have played pivotal roles in biotechnology as the prominent producers of enzymes,organic acids and antibiotics.Filamentous fungi are also important decomposers that contribute to the biological carbon cycle of plant biomass.Genetic engineering is a powerful approach for researchers not only to elucidate the gene function in filamentous fungi,but also to improve their production levels and minimize unwanted by-product formation.However,the efficiency of homologous integration in filamentous fungi is very low using classical genetic approaches.Due to the complicated growth and lifestyle of filamentous fungi,the development of genetic tools and gene editing is relatively slow,which hinders the basic research and biotechnological development of filamentous fungi.In recent years,genome editing technologies based on programmable nucleases have been developed as the powerful gene engineering tools in a wide variety of organisms.The most rapidly developed and wildly used technology is the class of RNA-guided Cas9 nuclease known as the adaptive immune system CRISPR(clustered regularly interspaced short palindromic repeats).The Cas9-sgRNA complex binds to the corresponding target site of the protospacer in a genome,and the specifically induces double-strand breaks.These breaks can be used as a basis for site-specific mutagenesis mediated by non-homologous end-joining or for the introduction of precise mutation or integration via homology-directed repair.CRISPR-Cas systems have recently enabled a wide range of applications for genome editing in many organisms.Remarkably,and in just the past few years,the CRISPR-Cas9 system has emerged as a more efficient strategy for gene editing in filamentous fungi.In this review,we describe the research progress of three most widely used genome editing systems,including the development of CRISPR-Cas technology and its applications in filamentous fungi,especially
作者
刘倩
李金根
张晨阳
李芳雅
田朝光
LIU Qian;LI Jingen;ZHANG Chenyang;LI Fangya;TIAN Chaoguang(Key Laboratory of Systems Microbial Biotechnology,Tianjin Institute of Industrial Biotechnology,Chinese Academy of Sciences,Tianjin 300308,China;National Technology Innovation Center of Synthetic Biology,Tianjin 300308,China;University of Chinese Academy of Sciences,Beijing 100049,China)
出处
《合成生物学》
CSCD
2021年第2期256-273,共18页
Synthetic Biology Journal
基金
国家重点研发计划“合成生物学”重点专项(2018FYA0900500)
天津市合成生物技术创新能力提升行动项目(TSBICIP-KJGG-006)
国家自然科学基金项目(31972878,31771386和31972879)
中国科学院青年创新促进会项目(2019180和2020183)。
