斑马鱼转基因平台的建立.pdfVIP

生物技术通报 ·研究报告· BIOTECHNOLOGY BULLETIN 2013年第10期 斑马鱼转基因平台的建立 刘丽丽  王健  王海胜  余凯敏  李国超  闫艳春 （中国农业科学院研究生院，北京 100081） 摘 要 ： 建立实验室斑马鱼转基因平台，旨在为实验室斑马鱼规模化繁殖、转基因斑马鱼研究提供技术流程、生物材料和 执行标准。以一对Tübingen 品系斑马鱼为材料，在实验室条件下进行扩大繁殖 ；将广谱表达绿色荧光蛋白GFP 真核载体pEGFP- N3 通过显微注射的手段转入斑马鱼胚胎，获得转基因斑马鱼；通过荧光显微镜观察斑马鱼GFP 表达情况，并初步筛选转基因个体； 以观察到GFP 表达的斑马鱼为材料，利用PCR 验证外源基因pEGFP-N3 在宿主细胞内的整合情况。结果显示，在实验室内建立了 完善的斑马鱼养殖系统，可以长期进行斑马鱼饲养、繁殖、育种等研究工作 ；建立了斑马鱼基础分子生物学试验体系，可以进行 斑马鱼分子水平的研究工作 ；建立了斑马鱼转基因平台，为构建转基因斑马鱼品系、外源基因整合机制等研究奠定基础。 关键词 ： 斑马鱼 繁殖 转基因平台 显微注射 Foundation of Platform for Zebrafish Transgenic Technology Liu Lili Wang Jian Wang Haisheng Yu Kaimin Li Guochao Yan Yanchun （ ， 100081 ） Graduate School of Chinese Academy of Agricultural Sciences Beijing Abstract:  The aim was to provide biomaterial, technique flow and protocol standard of transgenic zebrafish study via founding a platform of zebrafish transgenic technology. A couple of Tübingen zebrafish were multiplicated whose zygotes were microinjected with high active constitute plasmid pEGFP-N3. Injected eggs were collected and cultured and further observation. Fluorescent microscopic observation was followed by PCR on these embryos to screen and certify GFP expressed transgenic zebrafish. Results showed that the present study developed an intact cultivation system of zebrafish including its cultivation, reproduction and breeding. We also dug out the common emergency and proposed possible solutions during zebrafish maintenance. We designed the molecular processes with zebrafish embryo as material in our lab. Especially a platform of zebrafish transgenic technology was founded for the regeneration of transgenic