摘要
目的 :在酵母细胞表面展示登革 2型病毒 4 3株 (D2_4 3)的E基因 ,探索利用酵母表面展示系统建立DNA改组筛选平台的可行性。方法 :通过RT_PCR扩增获得D2_4 3的E基因 ,将该基因亚克隆至T载体后 ,再克隆至酵母表面展示载体pYD1,于酿酒酵母EBY10 0中利用半乳糖进行诱导表达。表达产物采用间接免疫荧光法和FACS进行检测。结果 :酵母表面展示产物可与D2_4 3的腹水抗体特异性地结合 ;在半乳糖诱导后 2 4h ,展示E蛋白的酵母细胞百分数达 2 2 .0 7%。结论 :本研究为建立基于酵母表面展示系统的DNA改组筛选平台奠定了基础。
Objective:To display the envelope protein of dengue virus type 2 strain 43 (D2-43) on yeast cell surface and explore a screening platform for DNA shuffling with yeast surface display system. Methods:The envelope gene of D2-43, which was amplified with RT-PCR, was subcloned into T vector, and then cloned into the yeast surface display vector pYD1.Epression was induced with galactose in Saccharomyces cerevisiae EBY100. Displayed products were detected with indirect immunofluorescence assay and FACS.Results:Displayed products on yeast cell surface interacted specifically with ascitic antibody against D2-43 and the percentage of yeast cells displaying the envelope protein was 22.07% after yeast cells were induced with galactose for 24 hours. Conclusion:The study lays a foundation to establish a screening platform for DNA shuffling based on yeast surface display system.
出处
《军事医学科学院院刊》
CSCD
北大核心
2004年第1期1-3,共3页
Bulletin of the Academy of Military Medical Sciences
基金
国家"8 63"计划资助项目 ( 2 0 0 1AA2 15 2 3 1)
