【摘要】 目的确定荧光蛋白标记是否影响GB1、GB1asa和GB2亚基在CHO细胞中的表达、亚细胞定位及功能。方法构建GB1-DsRed2和GB1asa-DsRed2真核表达载体,利用ELISA和IP3积累法检测融合蛋白在CHO细胞中的表达情况及功能,在激光扫描共聚焦显微镜下观测融合蛋白的亚细胞定位。结果荧光蛋白标记既不影响GB1、GB1asa和GB2亚基的表达,也不影响GB1和GB2在CHO细胞上的共定位过程,同时对二聚体的功能不存在显著的影响。结论GB1-DsRed2、GB1asa-DsRed2和GB2-EGFP在CHO细胞中具有正常的生理功能和分布,为进一步研究GABAB受体在细胞膜上的运动及分布以及各亚基之间的相互作用奠定了基础。更多还原

【Abstract】 Objective To study the expression,function and subcellular localization of different fluorescent protein tagged GB1-DsRed2,GB1asa-DsRed2 and GB2-EGFP in Chinese hamster ovary cell line CHO.Method The expression and function of the eukaryotic vector GB1-DsRed2 and GB1asa-DsRed2 were determined by ELISA and IP3 accumulation assay respectively,and the subcellular localization was conducted by laser scanning confocal microscope.Results The fusion protein GB1-DsRed2,GB1asa-DsRed2 and GB2-EGFP affected neither the expression nor the function of GABAB receptor,and the different fluorescent protein tagged GB1 and GB2 could also be co-localized in CHO cell line.Conclusion Fluorescent protein tagged GB1-DsRed2,GB1asa-DsRed2 and GB2-EGFP have the same functions and distribution patterns as wild types when expressing in CHO cells,and this finding provides a basis for further study on lateral movement and distribution of the GABAB receptor and the interaction between different subunits.更多还原