果蝇心脏发育标记基因Svp的多克隆抗体制备  

Preparation of Polyclonal Antibodies against Svp,the Marker Gene of Cardiac Development of Drosophila

作　　者：李玉玲 赵梦婧 李瑞可 张亚楠 吴秀山[1] 袁婺洲[1] LI Yuling;ZHAO Mengjing;LI Ruike;ZHANG Yanan;WU Xiushan;YUAN Wuzhou(The Center for Heart Development,the College of Life Sciences,Hunan Normal University,Changsha 410081,China)

机构地区：[1]湖南师范大学生命科学学院心脏发育研究中心,长沙410081

出　　处：《激光生物学报》2020年第5期433-437,445,共6页Acta Laser Biology Sinica

基　　金：国家自然科学基金项目(81370451);湖南省科技计划重点项目(2015DK3001)。

摘　　要：研究果蝇的心脏发育基因表达调控机制,其相应靶点的抗体必不可少。但目前市场上缺乏此类商用抗体,因此本研究选择果蝇心脏发育中的标记基因Svp(Seven-up),并采用DNA免疫技术来快速制备其抗体。首先通过聚合酶链式反应(PCR)扩增出Svp基因编码区部分序列,然后将其连接到真核表达载体pCAGGS-P7上,再将重组质粒pCAGGS-P7-Svp直接注射进小鼠体内,使外源基因在活体内表达,产生的抗原激活了小鼠的特异性细胞免疫和体液免疫,从而分泌相应的抗体,取小鼠的血清收集获得抗体。最后用蛋白质印迹(Western blot)和胚胎免疫荧光技术检测Svp抗体的效价和特异性。结果表明,制备的Svp多克隆抗体具有高效价和特异性,为后续的果蝇心脏发育研究奠定了基础。In order to study the regulation mechanism of gene expression in Drosophila heart development,obtaining the antibody of its corresponding target is essential,however so far there is no commercial antibodies used for research purpose,therefore we prepared antibodies rapidly by adopting DNA immune technology.Svp(seven-up)is one of the marker genes of cardiac development in Drosophila.In the first instance,Svp coding sequence which was obtained through PCR cloning was inserted into the pCAGGS-P7 eukaryotic expression vector.Secondly,we collected the antibodies by injecting recombinant vectors(pCAGGS-P7-Svp)directly into wild-type mice,making the exogenous genes expressed in the living body,through which antigens stimulated immune response and induces specific humoral and cellular immune response.Western blot and embryo immunofluorescence were used to detect the titer and specificity of Svp antibody.The result showed that polyclonal antibodies against Svp of high titer and specificity were obtained,which laid the foundation for further cardiac development research in Drosophila.

关 键 词：SVP 果蝇 心脏发育 DNA疫苗 多克隆抗体 

分 类 号：Q78[生物学—分子生物学]