用于胚胎活细胞显微镜观察的 SNAP 及 Halo 标记和染料引入方案。
SNAP- and Halo-tagging and dye introduction protocol for live microscopy in embryos.
机构信息
Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, MI 48109-1085, USA.
Department of Molecular, Cellular, and Developmental Biology, University of Michigan, Ann Arbor, MI 48109-1085, USA.
出版信息
STAR Protoc. 2022 Aug 18;3(3):101622. doi: 10.1016/j.xpro.2022.101622. eCollection 2022 Sep 16.
Traditional fluorescent proteins exhibit limitations in brightness and photostability that hinder optimal characterization of the dynamic cellular behavior of proteins of interest. SNAP- and Halo-tagging are alternatives to traditional fluorescent protein tagging utilizing bright, stable chemical dyes, which may improve signal-to-noise ratio. However, there has been limited use of this approach in developing organisms. Here, we present a protocol for implementing SNAP- and Halo-tagging in gastrula-stage embryos for live confocal microscopy. For complete details on the use and execution of this protocol, please refer to Varadarajan et al. (2022).
传统荧光蛋白在亮度和光稳定性方面存在局限性，这阻碍了对感兴趣的蛋白质的动态细胞行为进行最佳表征。SNAP 和 Halo 标记是利用明亮、稳定的化学染料替代传统荧光蛋白标记的方法，这可能会提高信噪比。然而，这种方法在发育生物中应用有限。在这里，我们提出了一种在原肠胚期胚胎中进行 SNAP 和 Halo 标记以进行活共聚焦显微镜的方案。有关此方案的使用和执行的完整详细信息，请参阅 Varadarajan 等人。（2022 年）。
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