Graduate School of Agriculture, Kyoto University, Kyoto 606-8502, Japan.
Institute of Evolutionary Biology (CSIC-Universitat Pompeu Fabra), Passeig Marítim de la Barceloneta 37, Barcelona 08003, Spain.
Cell Rep Methods. 2022 May 16;2(5):100215. doi: 10.1016/j.crmeth.2022.100215. eCollection 2022 May 23.
Current approaches for insect gene editing require microinjection of materials into early embryos. This severely limits the application of gene editing to a great number of insect species, especially to those whose reproduction systems preclude access to early embryos for injection. To overcome these limitations, we report a simple and accessible method for insect gene editing, termed "direct parental" CRISPR (DIPA-CRISPR). We show that injection of Cas9 ribonucleoproteins (RNPs) into the haemocoel of adult females efficiently introduces heritable mutations in developing oocytes. Importantly, commercially available standard Cas9 protein can be directly used for DIPA-CRISPR, which makes this approach highly practical and feasible. DIPA-CRISPR enables highly efficient gene editing in the cockroaches, on which conventional approaches cannot be applied, and in the model beetle . Due to its simplicity and accessibility, DIPA-CRISPR will greatly extend the application of gene editing technology to a wide variety of insects.
目前的昆虫基因编辑方法需要将材料微注射到早期胚胎中。这严重限制了基因编辑在大量昆虫物种中的应用，特别是对于那些生殖系统不允许注射早期胚胎的物种。为了克服这些限制，我们报告了一种简单易用的昆虫基因编辑方法，称为“直接亲本”CRISPR（DIPA-CRISPR）。我们表明，将 Cas9 核糖核蛋白（RNP）注射到成年雌性的血腔中，可以有效地在正在发育的卵母细胞中引入可遗传的突变。重要的是，商业上可获得的标准 Cas9 蛋白可以直接用于 DIPA-CRISPR，这使得该方法非常实用和可行。DIPA-CRISPR 使基因编辑在常规方法无法应用的蟑螂和模式甲虫中实现了高效。由于其简单性和易用性，DIPA-CRISPR 将极大地扩展基因编辑技术在各种昆虫中的应用。
