果蝇Bves蛋白的表达、纯化及多克隆抗体的制备  

The Expression,Purification of Bves Fusion Protein and Preparation of Its Polyclonal Antibody

作　　者：彭佩莹[1] 赵阳[1] 王静[1] 周英[1] 郝建军[1] 曾群[1] 陈思行[1] 任恋[1] 戴悦[1] 袁婺洲[1] 吴秀山[1] 邓云[1] 戴国[1] 

机构地区：[1]湖南师范大学蛋白质化学及鱼类发育生物学教育部重点实验室心脏发育研究中心,湖南长沙410081

出　　处：《中南医学科学杂志》2013年第3期221-224,246,共5页Medical Science Journal of Central South China

基　　金：国家自然科学基金(81170088);国家精品课程(教高函2009-21)

摘　　要：目的为了进一步研究细胞粘附分子Bves在肿瘤的发生及转移过程中的作用,需要获得Bves蛋白并制备其抗体。方法根据Flybase网站所提供的Bves基因序列,以果蝇的总RNA为模板进行PCR扩增,得到Bves部分编码序列,随后将其连接到pET-28a载体上获得原核表达载体。重组表达质粒经酶切测序鉴定后,转化入大肠杆菌BL21感受态细胞中,并用IPTG诱导融合蛋白的表达,使用活化的Ni-IDA凝胶柱亲和纯化,将纯化后得到的His-Bves融合蛋白免疫注射新西兰大白兔制备Bves多克隆抗体,并用Western blot对抗体效价进行检测。结果和结论获得了Bves原核表达重组融合蛋白及高效价的兔抗Bves多克隆抗体,为后期深入研究Bves基因的功能奠定了基础。Objective The Bves,as cell adhesion molecules,plays a certain role in tumorigenesis and metastasis.In order to further study the function of Bves,its protein and antibody must be obtained.Methods According to the Bves gene sequence provided on Flybase,the Drosophila RNA was constructed and then used for PCR amplification,part of the coding sequence was got,which could be connected to the prokaryotic expression vector pET-28a vector.The recombination plasmids were digested and sequenced correctly,and transformed into E.coli BL21 competent cells.The fusion protein was induced with IPTG,and purified by Ni-IDA gel column affinity.Then the purified His-Bves was immuned and injected to New Zealand white rabbits to prepare the polyclonal antibodies and its titers were detected by Western blot.Results and Conclusion The Bves prokaryotic expression recombination fusion protein and a high titer rabbit polyclonal antibody were gained.This study laid the foundation for later in-depth study for the function of Bves gene.

关 键 词：Bves 果蝇 融合蛋白 多克隆抗体 

分 类 号：Q785[生物学—分子生物学]