Transgenic fish as biosensors for superfund chemicals

转基因鱼将被用于研究农药对转基因报告基因表达的影响，这些报告基因报告了鱼类发育过程中雌激素反应、视黄酸反应通路和神经系统发育过程中的变化。我们的假设是：1)肝脏报告基因转基因鱼胚胎和幼虫可以作为生物传感器来检测环境毒物对整个生物体的影响。2)影响雌激素受体信号转导通路的化学物质可能存在独特的敏感性窗口，可以在鱼类模型中检测和评估。3)双双性报告基因将允许组织特异性表达研究和毒物对整个鱼胚胎的定量评估。4)为斑马鱼开发的转基因技术可用于本地物种，如鳉鱼、异交底鱼。本提案的具体目的是：1 .探索利用转基因斑马鱼模型筛选几种超级基金化学品，以检测GFP表达的差异。2。研究GFP阳性的原始生殖细胞进入发育中的生殖道的行为和结合情况，并观察在特定目的中研究的农药处理是否对鱼的性别决定有生物学影响。为适当的转基因启动子开发双双性载体，使荧光素酶基因（用于定量测量）和绿色荧光蛋白基因（用于信号的解剖定位）能够在配体诱导下表达。IV.根据具体目标I和II的结果，用最合适的转基因开发眼底模型。

Transgenic fish will be used to study the effect of pesticides upon gene expression of transgenic reporter genes that report changes in the estrogen-responsive, retinoic acid-responsive pathways in developing fish and in the development of the nervous system. Our hypothesis are: 1) Liver reporter transgenic fish embryos and larvae can be used as biosensors to detect whole organism effects of environmental toxicants. 2) There may be unique windows of sensitivity of chemicals which impact upon the estrogen receptor signal transduction pathway that can be detected and evaluated in fish models. 3) Dicistronic reporter genes will allow for both tissue specific expression studies plus quantitative evaluation of toxicants upon whole fish embryos. 4) Transgenic techniques developed for the zebrafish can be used for indigenous species such as the killifish, Fundulus heteroclitus. The specific aims of this proposal are: I. To explore the use of transgenic zebrafish models to screen several superfund chemicals for detectable differences of GFP expression. II. To examine the behavior and incorporation of the GFP positive primordial germ cells into the developing reproductive tract and to see if there are biological effects upon sexual determination of the fish through treatment with pesticides investigated in specific aim I. III. To develop dicistronic vectors for the appropriate transgenic promoters that will allow for ligand inducible expression of both a luciferase gene (for quantitative measurement) and a GFP gene (for anatomical localization of signal). IV. To develop Fundulus models with the most appropriate transgenes based upon the results of specific aims I and II.