Embryo Injections for CRISPR-Mediated Mutagenesis in the Ant Harpegnathos saltator
Embryo Injections for CRISPR-Mediated Mutagenesis in the Ant Harpegnathos saltator
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DOI：
10.3791/61930
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发表时间：
2021-02-01
影响因子：
1.2
通讯作者：
Yan, Hua
中科院分区：
文献类型：
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作者：
Sieber, Kayli;Saar, Maya;Yan, Hua
真社会性昆虫的社会行为和生殖分工等独特特征是由遗传系统控制的。为了解决基因如何调节社会特征，我们通过将CRISPR复合物递送到合胞期的年轻胚胎中来开发突变蚂蚁。在这里，我们提供了一个CRISPR介导的诱变Harpegnathos saltator的方案，Harpegnathos saltator是一种表现出惊人的表型可塑性的重要物种。H.跳蚁很容易在实验室环境中饲养。收集胚胎，使用自制的石英针用Cas9蛋白和体外合成的小向导RNA（sgRNA）进行显微注射。注射后的胚胎在殖民地外饲养。在第一只幼虫出现后，所有的胚胎和幼虫都被运送到一个有几个保育员的巢箱中进行进一步的发育。该方案适用于诱导突变，用于分析蚂蚁的种姓特异性生理和社会行为，但也可应用于更广泛的hyperterans和其他昆虫。
The unique traits of eusocial insects, such as social behavior and reproductive division of labor, are controlled by their genetic system. To address how genes regulate social traits, we have developed mutant ants via delivery of CRISPR complex into young embryos during their syncytial stage. Here, we provide a protocol of CRISPR-mediated mutagenesis in Harpegnathos saltator, a ponerine ant species that displays striking phenotypic plasticity. H. saltator ants are readily reared in a laboratory setting. Embryos are collected for microinjection with Cas9 proteins and in vitro synthesized small guide RNAs (sgRNAs) using home-made quartz needles. Post-injection embryos are reared outside the colony. Following emergence of the first larva, all embryos and larvae are transported to a nest box with a few nursing workers for further development. This protocol is suitable for inducing mutagenesis for analysis of caste-specific physiology and social behavior in ants, but may also be applied to a broader spectrum of hymenopterans and other insects.