Kim Minyoung, Hutchins Erica J
Department of Cell and Tissue Biology, University of California San Francisco, San Francisco, CA, USA; Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California San Francisco, San Francisco, CA, USA; Oral and Craniofacial Sciences Graduate Program, School of Dentistry, University of California San Francisco, San Francisco, CA, USA.
Department of Cell and Tissue Biology, University of California San Francisco, San Francisco, CA, USA; Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California San Francisco, San Francisco, CA, USA; Oral and Craniofacial Sciences Graduate Program, School of Dentistry, University of California San Francisco, San Francisco, CA, USA.
Dev Biol. 2025 Mar;519:5-12. doi: 10.1016/j.ydbio.2024.11.013. Epub 2024 Nov 30.
The chick embryo is a classical model system commonly used in developmental biology due to its amenability to gene perturbation experiments. Pairing this powerful model organism with cutting-edge technology can significantly expand the range of experiments that can be performed. Recently, the CRISPR-Cas13d system has been successfully adapted for use in zebrafish, medaka, killifish, and mouse embryos to achieve targeted gene expression knockdown. Despite its success in other animal models, no prior study has explored the potential of CRISPR-Cas13d in the chick. Here, we present an adaptation of the CRISPR-Cas13d system to achieve targeted gene expression knockdown in the chick embryo. As proof-of-principle, we demonstrate the knockdown of PAX7, an early neural crest marker. Application of this adapted CRISPR-Cas13d technique resulted in effective knockdown of PAX7 expression and function, comparable to knockdown achieved by translation-blocking morpholino. CRISPR-Cas13d complements preexisting knockdown tools such as CRISPR-Cas9 and morpholinos, thereby expanding the experimental potential and versatility of the chick model system.
鸡胚是发育生物学中常用的经典模型系统，因为它适合进行基因干扰实验。将这种强大的模式生物与前沿技术相结合，可以显著扩展可进行的实验范围。最近，CRISPR-Cas13d系统已成功应用于斑马鱼、青鳉、鳉鱼和小鼠胚胎，以实现靶向基因表达敲低。尽管它在其他动物模型中取得了成功，但之前没有研究探索过CRISPR-Cas13d在鸡中的潜力。在这里，我们展示了一种对CRISPR-Cas13d系统的改进，以在鸡胚中实现靶向基因表达敲低。作为原理验证，我们证明了早期神经嵴标记物PAX7的敲低。应用这种改进的CRISPR-Cas13d技术导致PAX7表达和功能的有效敲低，与通过翻译阻断吗啉代实现的敲低效果相当。CRISPR-Cas13d补充了现有的敲低工具，如CRISPR-Cas9和吗啉代，从而扩展了鸡模型系统的实验潜力和通用性。
