Gene transfection is an indispensable approach for studying gene function since it provides important information on gain-and/or loss-of-function.Chick embryos are also extensively employed for studying biological fun...Gene transfection is an indispensable approach for studying gene function since it provides important information on gain-and/or loss-of-function.Chick embryos are also extensively employed for studying biological function since they are easily accessible and can be maintained alive after manipulation.The combination of both techniques presents a powerful approach to understanding how genes regulate embryo development.Furthermore,combining these approaches with tissue transplant techniques make even more attractive for elucidate gene function.Electroporation,employing parallelly fashioned electrodes,has been widely used in chick embryos.However,experimenters have been frustrated by unsuccessfully transfection in some embryonic tissue of interest because the electrodes were improperly positioned.We presently demonstrated the different patterns of organizing and positioning the electrodes,in combination with tissue transplantation,to efficiently and specifically transfect the chick embryonic head,trunk neural tube,heart tube,somites and neural crest cells with the GFP reporter gene.展开更多
目的借助生物物理技术建立一种高效实用的脑内基因转染技术,探讨其在神经科学研究中的应用。方法选取孕15 d C57系小鼠,取出子宫角,用自制玻璃电极将目的质粒和荧光质粒混匀后注入胎鼠侧脑室,应用BTX电转染仪在胎鼠脑两颞极间给予设定...目的借助生物物理技术建立一种高效实用的脑内基因转染技术,探讨其在神经科学研究中的应用。方法选取孕15 d C57系小鼠,取出子宫角,用自制玻璃电极将目的质粒和荧光质粒混匀后注入胎鼠侧脑室,应用BTX电转染仪在胎鼠脑两颞极间给予设定的电脉冲刺激,后将胎鼠放入孕鼠腹腔让其自然出生后8 d取脑,冰冻切片,抗GFP、抗Tubulin免疫组化染色,荧光显微镜下观察。结果在小鼠大脑皮层Ⅱ、Ⅲ层,转染CAG-EGFP的脑片可见绿色荧光细胞带,转染细胞抗GFP免疫组化染色阳性;转染DS-red的呈红色荧光细胞带,转染细胞均呈抗Tubulin免疫组化染色阳性。结论子宫内胚胎电转染技术是一个很好的在体基因转染技术,可为神经科学研究提供一个很好的技术平台。展开更多
基金supported by the National Natural Science Foundation of China(31071054,30971493,31271455)the National Basic Research Program of China(2010CB529703)the Natural Science Foundation of Guangdong Province(S2011010001593)
文摘Gene transfection is an indispensable approach for studying gene function since it provides important information on gain-and/or loss-of-function.Chick embryos are also extensively employed for studying biological function since they are easily accessible and can be maintained alive after manipulation.The combination of both techniques presents a powerful approach to understanding how genes regulate embryo development.Furthermore,combining these approaches with tissue transplant techniques make even more attractive for elucidate gene function.Electroporation,employing parallelly fashioned electrodes,has been widely used in chick embryos.However,experimenters have been frustrated by unsuccessfully transfection in some embryonic tissue of interest because the electrodes were improperly positioned.We presently demonstrated the different patterns of organizing and positioning the electrodes,in combination with tissue transplantation,to efficiently and specifically transfect the chick embryonic head,trunk neural tube,heart tube,somites and neural crest cells with the GFP reporter gene.
文摘目的借助生物物理技术建立一种高效实用的脑内基因转染技术,探讨其在神经科学研究中的应用。方法选取孕15 d C57系小鼠,取出子宫角,用自制玻璃电极将目的质粒和荧光质粒混匀后注入胎鼠侧脑室,应用BTX电转染仪在胎鼠脑两颞极间给予设定的电脉冲刺激,后将胎鼠放入孕鼠腹腔让其自然出生后8 d取脑,冰冻切片,抗GFP、抗Tubulin免疫组化染色,荧光显微镜下观察。结果在小鼠大脑皮层Ⅱ、Ⅲ层,转染CAG-EGFP的脑片可见绿色荧光细胞带,转染细胞抗GFP免疫组化染色阳性;转染DS-red的呈红色荧光细胞带,转染细胞均呈抗Tubulin免疫组化染色阳性。结论子宫内胚胎电转染技术是一个很好的在体基因转染技术,可为神经科学研究提供一个很好的技术平台。
