信号肽对鸡Igλ轻链绿色荧光蛋白重组分子在COS7细胞上分泌表达的影响  

Effect of Signal Peptides on the Secretable Expression of Recombinant Chicken Igλ/GFP in COS7 Cells

作　　者：张红[1] 王健[1] 李姣[2] 易明林[2] 韩鸿鹏[1] 张改平[2] 

机构地区：[1]河南教育学院,河南郑州450003 [2]河南省动物免疫学重点实验室,河南郑州450002

出　　处：《河南农业科学》2008年第2期99-102,共4页Journal of Henan Agricultural Sciences

基　　金：国家自然科学基金项目(30571388);河南省基础与前沿项目(072300430110);河南教育学院重点学科资助

摘　　要：分别将鸡Igλ轻链信号肽、小鼠纤溶酶原信号肽与pEGFP-C1载体的绿色荧光蛋白N端融合,产生带有信号肽的中间载体pEGFP-SPc和pEGFP-SPm。对鸡Igλ轻链基因的定向克隆,构建了带有纯化标签的鸡Igλ轻链绿色荧光蛋白真核表达载体pEGFP-SPc-λ和pEGFP-SPm-λ。转染COS7细胞后,荧光显微镜观察及Western blotting检测融合蛋白的分泌表达,结果显示,鸡Igλ轻链信号肽能引导鸡Igλ轻链绿色荧光蛋白融合分子的分泌表达;而小鼠纤溶酶原信号肽不具备引导该重组蛋白分泌表达的功能。表明信号肽在蛋白分泌表达中的关键作用,重组蛋白的分泌表达要选择合适的信号肽。The signal peptides of chicken Igλ light chain and murine plasmingen were fused to the GFP N terminus using site-directed mutagenesis to construct the vectors with signal sequence, pEGFP-SPc and pEGFP-SPm. The chicken Igλ gene was inserted into the two vectors with signal sequence after double enzyme cutting. The results revealed that the vectors, pEGFP-SPc-λ and pEGFP-SPm-λ ,with fusion gene of chicken Igλ and green fluroscent protein （GFP） were successfully constructed. The constructed recombinant vectors were transiently transfected into COS7 cells via lipofectamin. Secretable expression of Igλ/GFP in COS7 cells transfected with vector pEGFP-SPc-λ was observed under fluroscent microscope and in Western blotting. Expression of GFP fusion protein was located principally in plasma of COS7 cells transfected with pEGFP- SPm-λ vector. These findings suggest that signal peptides play a key role in secreted protein translocation. Signal peptide needs to be carefully chosen to guarantee the secretable expression of recombinant proteins.

关 键 词：鸡Igλ轻链信号肽 小鼠纤溶酶原信号肽 绿色荧光蛋白 分泌表达 

分 类 号：Q78[生物学—分子生物学]