Microinjection for precision genome editing in Caenorhabditis elegans.
Microinjection for precision genome editing in Caenorhabditis elegans.
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DOI：
10.1016/j.xpro.2021.100748
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发表时间：
2021-09-17
期刊：
影响因子：
--
通讯作者：
Mello CC
中科院分区：
文献类型：
--
作者：
Ghanta KS;Ishidate T;Mello CC
在秀丽隐杆线虫中，靶向基因组编辑技术现在通常用于生成种系编辑。秀丽隐杆线虫生殖系编辑的显著容易性归因于粗线期卵巢的合胞体性质，其易于通过显微注射进入。该方案描述了整个CRISPR-Cas基因组编辑程序的分步细节和故障排除提示，包括gRNA设计和核糖核蛋白复合物的显微注射，然后在秀丽隐杆线虫中进行筛选和基因分型，以帮助访问这个强大的遗传动物系统。有关本方案使用和执行的完整详细信息，请参阅。在秀丽隐杆线虫中进行CRISPR-Cas基因组编辑的分步显微注射方案解决显微注射和基因组编辑方案问题的提示熔化dsDNA供体分子增强精确的基因组编辑单个注射的雌雄同体可以产生数十个精确编辑的后代在秀丽隐杆线虫中，靶向基因组编辑技术现在通常用于生成种系编辑。秀丽隐杆线虫生殖系编辑的显著容易性归因于粗线期卵巢的合胞体性质，其易于通过显微注射进入。该方案描述了整个CRISPR-Cas基因组编辑程序的分步细节和故障排除提示，包括gRNA设计和核糖核蛋白复合物的显微注射，然后在秀丽隐杆线虫中进行筛选和基因分型，以帮助访问这个强大的遗传动物系统。
In Caenorhabditis elegans, targeted genome editing techniques are now routinely used to generate germline edits. The remarkable ease of C. elegans germline editing is attributed to the syncytial nature of the pachytene ovary which is easily accessed by microinjection. This protocol describes the step-by-step details and troubleshooting tips for the entire CRISPR-Cas genome editing procedure, including gRNA design and microinjection of ribonucleoprotein complexes, followed by screening and genotyping in C. elegans, to help accessing this powerful genetic animal system. For complete details on the use and execution of this protocol, please refer to. Step-by-step microinjection protocol for CRISPR-Cas genome editing in C. elegans Tips to troubleshoot microinjection and genome editing protocols Melting dsDNA donor molecules potentiates precision genome editing A single injected hermaphrodite can produce dozens of precisely edited progenies In Caenorhabditis elegans, targeted genome editing techniques are now routinely used to generate germline edits. The remarkable ease of C. elegans germline editing is attributed to the syncytial nature of the pachytene ovary which is easily accessed by microinjection. This protocol describes the step-by-step details and troubleshooting tips for the entire CRISPR-Cas genome editing procedure, including gRNA design and microinjection of ribonucleoprotein complexes, followed by screening and genotyping in C. elegans, to help accessing this powerful genetic animal system.
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