Rap1基因在斑马鱼胚胎早期发育过程中的时空表达谱研究  

Spatiotemporal expression spectrum of Rap1 gene in zebrafish early development process

作　　者：杨小燕[1] 何志旭[1,2] 舒莉萍[2,3] 金皎[1] 黄璟[1] 吴莎莎[1] 马健娟[1] 

机构地区：[1]贵州医科大学附属医院儿科,550004 [2]贵州医科大学组织工程与干细胞实验中心,550004 [3]贵州医科大学免疫学教研室,贵阳550004

出　　处：《重庆医学》2016年第20期2748-2751,共4页Chongqing medicine

基　　金：国家自然科学基金面上项目(30960412);贵州省科技基础条件平台项目([2009]4005);贵州省优秀教育人才省长专项资金($2008-4)

摘　　要：目的选择斑马鱼作为实验动物模型，研究Rap1基因在胚胎早期发育过程中的时空表达规律。方法从斑马鱼胚胎cDNA中克隆Rap1基因片段，将Rap1基因片段和pCS2＋质粒进行体外连接重组，重组质粒经双酶切、菌落聚合酶链反应（PCR）及测序鉴定正确后，经T3RNA体外转录体系合成DIG标记的Rap1基因反义mRNA探针，采用整胚原位杂交方法检测Rap1在斑马鱼胚胎早期发育过程的表达情况。结果在斑马鱼胚胎0．75hpf的细胞分裂连接处、3．70hpf和6．00hpf的动物极、12．00～72．00hpf的脊索神经部位均可见Rap1基因的阳性杂交信号。结论Rap1基因在斑马鱼脊索神经系统的早期发育过程中可能起到重要的调控作用。Objective To choose zebrafish as the experimental animal model for studying the spatiotemporal expression rule of rapl gen in zebrafish embryo early development process. Methods The Rap1 gene fragment was cloned from the zebrafisb embyoscDNA,then the Rap1 gene fragment and pCS2＋ plasmid were performed the in vitro connetion and recombination was extracted, the combinant plasmid was correct after the double enzyme digestion,colony PCR and sequencing identification. T3 RNA polymerase in vitro transcription system was used to obtain the digoxin（DIG）-labeled anti-sense mRNA probe of Rapl gene. The whole mount in situ hybridization method was adopted to detect the Rapl expression in zebrafish embryo early development process. Results The positive hybridization signal of Rap1 gene was detected at the cell division junction region of 0.75 hpf,animal pole of 3.70 hpf and 6.00 hpf, and notochord of 12.00- 72.00 hpf. Conclusion Rapl gene might be involved in the early development process of notochord nervous system in zebrafish.

关 键 词：Rap1基因 斑马鱼 原位杂交 基因表达 

分 类 号：Q344[生物学—遗传学]