HIV-1 gp41的酵母表面展示及表达优化

被引:1
作者
向柱方 [1 ]
林影 [1 ]
叶波 [1 ]
韩双艳 [1 ]
赵树进 [2 ]
机构
[1] 华南理工大学生物科学与工程学院
[2] 广州军区广州总医院
基金
广东省科技计划;
关键词
酵母展示技术; HIV-1 gp41; 免疫荧光染色; 流式细胞仪;
D O I
10.13345/j.cjb.2008.04.016
中图分类号
Q78 [基因工程(遗传工程)];
学科分类号
071007 ; 0836 ; 090102 ;
摘要
以His标签检测蛋白的表达,利用酿酒酵母表面展示系统,成功地将HIV-1 gp41片段锚定在酵母表面,并检测到gp41的活性。以pMD18T-gp41为模板,通过PCR技术克隆了gp41基因,将gp41基因通过双酶切连接到载体pICAS-His上,构建了gp41酵母表面展示载体,并将其转化至酿酒酵母(Saccharomyces cerevisiae)MT8-1中。重组菌经培养,利用免疫荧光染色方法进行染色,显微镜观察发现重组酵母细胞表面有绿色荧光,流式细胞仪结果进一步证实gp41正确折叠展示于酵母细胞表面。采用不同浓度的葡萄糖培养基进行表达优化。当葡萄糖浓度为1%时,82.46%的酵母细胞表达了gp41抗原;随着葡萄糖浓度升高,蛋白表达受到抑制。
引用
收藏
页码:684 / 689
页数:6
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