The olive fruit fly, Bactrocera oleae (Diptera: Tephritidae), is the most destructive insect pest of olive cultivation, causing significant economic and production losses. Here, we present the establishment of the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 methodology for gene disruption in this species. We performed targeted mutagenesis of the autosomal gene white (Bo‐we), by injecting into early embryos in vitro preassembled and solubilized Cas9 ribonucleoprotein complexes loaded with two gene‐specific single‐guide RNAs. Gene disruption of Bo‐we led to somatic mosaicism of the adult eye color. Large eye patches or even an entire eye lost the iridescent reddish color, indicating the successful biallelic mutagenesis in somatic cells. Cas9 induced either indels in each of the two simultaneously targeted Bo‐we sites or a large deletion of the intervening region. This study demonstrates the first efficient implementation of the CRISPR/Cas9 technology in the olive fly, providing new opportunities towards the development of novel genetic tools for its control.

中文翻译：

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通过CRISPR / Cas9核果蛋白复合物在橄榄果蝇Bactrocera oleae中进行定向诱变。

橄榄果蝇Bactrocera oleae（Diptera：Tephritidae）是橄榄栽培中最具破坏性的害虫，造成重大的经济和生产损失。在这里，我们介绍了该物种中基因破坏的簇状规则间隔的短回文重复序列（CRISPR）/ Cas9方法的建立。我们通过将预先组装并溶解了两个基因特异性单向导RNA的Cas9核糖核蛋白复合物注射到体外早期胚中，对常染色体基因白色（Bo-we）进行了定向诱变。Bo-we的基因破坏导致了成年人眼睛颜色的体细胞镶嵌。大眼斑甚至整个眼睛失去了虹彩带红色，表明在体细胞中成功进行了双等位基因诱变。Cas9会在两个同时定位的Bo-we位点中的每一个中诱导插入缺失，或在中间区域有较大的缺失。这项研究证明了CRISPR / Cas9技术在橄榄蝇中的首次有效实施，为开发控制其的新型遗传工具提供了新的机会。