袁燕蒙,葛玉丹,张舟
YUAN Yan-meng,GE Yu-dan,ZHANG Zhou

• 1:沈阳药科大学生命科学与生物制药学院

摘要(Abstract)：

目的利用分子生物学方法构建大鼠谷氨酰胺转运蛋白1重组质粒p EGFP-N1-SNAT1并进行鉴定。方法对载体p EGFP-N1和质粒p BK-CMV(Δ[1098-1300])-SNAT1双酶切,纯化后连接,构建重组质粒p EGFP-N1-SNAT1。用Western blot检测融合蛋白的表达,采用免疫荧光检测SNAT1在细胞膜上的表达和定位。结果成功构建重组质粒p EGFP-N1-SNAT1并正常表达、定位于细胞膜上。结论重组质粒p EGFP-N1-SNAT1的成功构建为研究SNAT1的结构和功能提供了有效工具。
Objective To determine the expression and localization of SNAT1,an eukaryotic expression plasmid with EGFP tag at the C terminus of SNAT1 was constructed successfully. Methods Methods The vector p EGFP-N1 and plasmid p BK-CMV( Δ[1098-1300])-SNAT1 were digested with Sac II and Hind III separately,and then the DNA fragments were purified and ligated. After constructing the recombinant plasmid p EGFP-N1-SNAT1,the expression and cell membrane localization of SNAT1 were detected by Western blot using anti-GFP antibody and laser scanning confocal microscope. Results The recombinant plasmid p EGFP-N1-SNAT1 was successfully constructed and identified. Conclusions Successful construction of the recombinant plasmid p EGFP-N1-SNAT1 provides an effective tool for studying structure and function of SNAT1 in the future.

关键词(KeyWords)： 谷氨酰胺转运蛋白SNAT1;融合蛋白;增强型绿色荧光蛋白
glutamine transporters SNAT1;fusion protein;enhanced green fluorescence protein(EGFP)

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金资助项目(31270883)

作者(Author): 袁燕蒙,葛玉丹,张舟
YUAN Yan-meng,GE Yu-dan,ZHANG Zhou

DOI: 10.14066/j.cnki.cn21-1349/r.2017.05.012

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