重组Zonadhesin蛋白D4结构域对小鼠体外受精的影响  

Effect of recombinant zonadhesin protein D4 domain on in vitro fertilization of mouse

作　　者：吕正梅[1,2] 王旻[1] 胡燕琴[1] 郭强苏[1] 徐晨[1] 

机构地区：[1]上海交通大学医学院组织学与胚胎学教研室上海市生殖医学重点实验室,上海200025 [2]安徽医科大学组织学与胚胎学教研室,合肥230032

出　　处：《解剖学杂志》2009年第2期140-143,共4页Chinese Journal of Anatomy

基　　金：国家自然科学基金(30470909)

摘　　要：目的:研究Zonadhesin蛋白D4domain在受精过程中的作用。方法:将小鼠Zonadhesin D4结构域分亚结构域(VWDsubdomain及余C-端片段)克隆,在E.coli诱导表达目的蛋白,将纯化重组蛋白片段为抗原,制备兔抗鼠D4结构域多克隆抗体,免疫印迹法、精子间接免疫荧光染色鉴定多克隆抗体的特异性及D4结构域在小鼠精子表达。用多抗处理获能小鼠精子,然后与小鼠超排卵子进行体外受精及精子-透明带结合实验。结果:D4亚结构域成功克隆表达,制备了能特异性识别精子天然Zonadhesin D4的多克隆抗体,多抗处理组精子与卵透明带结合数与对照组差异无统计学意义,卵子的受精率也无明显差异。结论:抗重组Zonadhesin D4结构域的多抗对小鼠体外受精无抑制作用。Objective： To investigate the possible functions of zonadhesin D4 domain in the process of fertilization. Methods： Two cDNA fragments coding for VWD subdomain and the rest C-termianl fragment of D4 domain were cloned and expressed in E. coil Antibodies against recombinant D4 domain were raised in New Zealand white rabbits immunized with recombinant fragments. The expression of this protein on mouse sperm and the specificity of the polyclonal antibodies were determined by indirect immunofluorescent staining and Western blot analysis. After treated sperm with these antibodies, sperm-zona pellucida binding assay and in vitro fertilization were carried out in BALB/c mice. Results： Recombinant D4 domain was successfully cloned and expressed. The expression of the protein was limited on the anterior head of mouse sperm and the antibodies against it were specific. The treatment of sperm with polyclonal antibodies did not significantly decrease the average number of sperm bound to zona pellucida. Meanwhile, the percentage of fertilized eggs was not decreased significantly in comparison to control rabbit IgG. Conclusion： The results suggest that polyclonal antibodies against recombinant zonadhesin D4 domain may not inhibit in vitro fertilization of mouse.

关 键 词：Zonadhesin 多克隆抗体 体外受精 小鼠 

分 类 号：Q813[生物学—生物工程]