A nickase Cas9 gene-drive system promotes super-Mendelian inheritance in Drosophila.-猫眼课题宝

DOI：

10.1016/j.celrep.2022.110843

发表时间：

2022-05-24

期刊：

CELL REPORTS

影响因子：

8.8

通讯作者：

中科院分区：

生物学1区

文献类型：

作者：

Del Amo, Victor Lopez;Juste, Sara Sanz;Gantz, Valentino M.

来源链接：

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9190248/

基于CRISPR的基因驱动已被提议用于管理昆虫种群，包括传播疾病的蚊子，因为它们能够使其遗传偏向于超孟德尔率（>50%）。目前的技术使用Cas9，其将DNA双链断裂引入相对的野生型等位基因中，以通过DNA同源性定向修复用基因驱动等位基因的拷贝替换它。然而，不同Cas9版本的使用尚未探索，替代方法可以增加基因驱动设计的可用工具包。在这里，我们报告了一种依赖于Cas9切口酶的基因驱动，Cas9切口酶在DNA中产生交错的成对切口，以繁殖工程化的基因驱动盒。我们表明，在系统中产生5′突出端产生有效的等位基因转换。切口酶基因驱动排列产生大的、定型的缺失，这有利于在靶向必需基因时消除携带小突变的存活动物。我们的切口酶方法应该扩大基因驱动安排的范围，旨在应用于蚊子和其他动物。使用野生型Cas9的基因驱动提供了对抗媒介传播疾病的解决方案，但需要替代策略来增加可用的工具包。López Del Amo等人描述了一种基于切口酶的基因驱动系统，该系统促进果蝇种系中工程化盒的超孟德尔遗传，为基于CRISPR的基因驱动提供了替代设计。

CRISPR-based gene-drives have been proposed for managing insect populations, including disease-transmitting mosquitoes, due to their ability to bias their inheritance toward super-Mendelian rates (>50%). Current technologies use a Cas9 that introduces DNA double-strand breaks into the opposing wild-type allele to replace it with a copy of the gene-drive allele via DNA homology-directed repair. However, the use of different Cas9 versions is unexplored, and alternative approaches could increase the available toolkit for gene-drive designs. Here, we report a gene-drive that relies on Cas9 nickases that generate staggered paired nicks in DNA to propagate the engineered gene-drive cassette. We show that generating 5′ overhangs in the system yields efficient allelic conversion. The nickase gene-drive arrangement produces large, stereotyped deletions that are advantageous to eliminate viable animals carrying small mutations when targeting essential genes. Our nickase approach should expand the repertoire for gene-drive arrangements aimed at applications in mosquitoes and beyond. Gene-drives using wild-type Cas9 offer solutions to fight vector-borne diseases, yet alternative strategies are needed to increase the available toolkit. López Del Amo et al. describe a nickase-based gene-drive system that promotes super-Mendelian inheritance of an engineered cassette in the Drosophila germ line, providing alternative designs for CRISPR-based gene-drive.