Nakamoto Masaru, Nakamoto Chizu
Department of Biology, Neils Science Center & Center for the Sciences, Valparaiso University, Valparaiso, IN, USA.
Methods Mol Biol. 2020;2092:91-108. doi: 10.1007/978-1-0716-0175-4_8.
The embryonic chick has long been a favorite model system for in vivo studies of vertebrate development. However, a major technical limitation of the chick embryo has been the lack of efficient loss-of-function approaches for analyses of gene functions. Here, we describe a methodology in which a transgene encoding artificial microRNA sequences is introduced into embryonic chick retinal cells by in ovo electroporation and integrated into the genome using the Tol2 transposon system. We show that this methodology can induce potent and stable suppression of gene expression. This technique therefore provides a rapid and robust loss-of-function approach for studies of gene function in the developing retina.
长期以来,胚胎期的鸡一直是脊椎动物发育体内研究中备受青睐的模型系统。然而,鸡胚的一个主要技术局限在于缺乏用于基因功能分析的高效功能缺失方法。在此,我们描述了一种方法,即通过胚内电穿孔将编码人工微小RNA序列的转基因导入胚胎期鸡的视网膜细胞,并使用Tol2转座子系统将其整合到基因组中。我们证明,该方法能够有效且稳定地抑制基因表达。因此,这项技术为发育中的视网膜基因功能研究提供了一种快速且可靠的功能缺失方法。