CRISPR/Cas9系统递送方式的发展及其在昆虫研究中的应用

Development of CRISPR/Cas9 System Delivery Methods and Their Applications in Insect Research

CRISPR/Cas9基因编辑系统自问世以来,极大地推进了人类对生物基因功能的认识。CRISPR/Cas9通常采用显微注射昆虫胚胎的途径来递送基因编辑试剂,由于昆虫具有多种生殖方式,因此对于某些昆虫种类如具有坚硬卵鞘的蟑螂和胎生的蚜虫等,应用传统的CRISPR/Cas9系统的递送方式来了解其基因功能就难以实现。目前,在传统的CRISPR/Cas9递送方式上发展出来的不需要注射昆虫早期单细胞胚胎的方法已有相关报道:ReMOT (receptor-mediated ovary transduction)和DIPA-CRISPR (“direct parental”CRISPR),它们可以将编辑试剂直接注射到成年雌性个体的血腔中,能够有效地在发育中的卵母细胞中引入遗传突变,同时简化了基因编辑的过程,这类方法已经成功应用于埃及伊蚊、德国小蠊和赤拟谷盗等昆虫的研究。本文对CRISPR/Cas9系统的结构、作用机制、传统递送方式及其发展出的ReMOT和DIPA-CRISPR递送方法在昆虫研究中的应用现状与前景进行了综述,以期为更多物种的基因编辑研究提供参考。

Since the emergence of the CRISPR/Cas9 gene editing system,there has been significant advancement in understanding biological gene functions.CRISPR/Cas9 is typically delivered via microinjection into insect embryos for gene editing.However,due to diverse reproductive modes in insects,it is difficult to use traditional CRISPR/Cas9 delivery methods to study the gene functions of some insect species,such as cockroaches with rigid egg cases and viviparous aphids.In the current CRISPR/Cas9 technology,alternative delivery methods have been developed,which do not require injection into early-stage insect embryos and have been reported as ReMOT(receptor-mediated ovary transduction)and DIPA-CRISPR(“direct parental”CRISPR).These methods allow the direct injection of editing reagents into the hemocoel of adult female individuals,introducing genetic mutations effectively into developing oocytes and simplifying the gene editing process.They have been successfully applied to the study of insects such as Aedes aegypti,Blattella germanica,and Tribolium castaneum.This article reviews the structure,mechanism,and traditional delivery methods of the CRISPR/Cas9 system,as well as the application and prospects of the newly developed Re-MOT and DIPA-CRISPR delivery methods in insect research,aiming to provide guidance for gene editing in more species.