活体电转化技术在新生小鼠视网膜中的应用

活体电转化技术是在高电压的脉冲作用下,瞬态增加细胞膜的渗透性从而将外源基因高效导入细胞的方法。与病毒载体等其他方法相比,活体电转化技术具有安全、高效、快速、稳定及应用范围广等优点,近年来在很多组织和器官中得到广泛使用,包括在眼科研究领域。文章介绍了活体电转化技术在新生小鼠视网膜中的应用,通过新生小鼠视网膜下注射的方法,经几次高电压的脉冲,将高浓度的绿色荧光蛋白表达质粒导入新生小鼠视网膜细胞内。通过冰冻切片观察绿色荧光蛋白在视网膜中的表达。结果表明绿色荧光蛋白在视网膜外核层高表达,证实了活体电转化技术可以将外源基因高效、快捷的导入视网膜,从而为研究视网膜发育及功能提供一种有效的手段。

In vivo electroporation of newborn mouse retina

In vivo electroporation is the process referred to a transient increase in the permeability of cell membranes upon high electric field pulses and delivery of engineered constructs into cells. Compared with the viral-mediated gene transfer system, the in vivo electroporation technique has several advantages, such as safe, high efficiency, rapid, stable and wide application. Thus, this technique has been widely used in the studies of many tissues or organs, including the eye. Here, we report the application of in vivo electroporation in the newborn mouse retina. DNA plasmid of GFP expression vector with high concentration was directly injected into the subretinal space of neonatal mouse pups. The DNA was then transfected into the retinal cells after several pulses of high voltage. Transfected GFP allowed clear visualization of cell morphologies in cryo-sections and the GFP was highly expressed in retinal outer nuclear layer. The results showed that this technique can effectively transfer the genes into retinal cells. In vivo electroporation will be a useful tool for the study of retinal development and function.