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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 construct and identify the recombinant plasmid p EGFP-N1-SNAT1 of rat glutamine transporter 1 by molecular biology method. Methods The vector p EGFP-N1 and plasmid p BK-CMV (Δ [1098-1300]) - SNAT1 were double-digested and purified. The recombinant plasmid p EGFP-N1-SNAT1 was constructed. The expression of fusion protein was detected by Western blot and the expression and localization of SNAT1 on the cell membrane was detected by immunofluorescence. Results The recombinant plasmid p EGFP-N1-SNAT1 was successfully constructed and expressed on the cell membrane. Conclusion The successful construction of recombinant plasmid p EGFP-N1-SNAT1 provides an effective tool for studying the structure and function of SNAT1.