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和神经系统发育中雌激素反应和维甲酸反应途径的变化。我们的假设是：1)肝脏报告基因转基因鱼胚胎和仔鱼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)在FISH模型中，可能存在影响雌激素受体信号转导途径的化学物质的独特敏感性窗口。3)双顺反子报告基因将允许进行组织特异性表达研究以及对整个鱼类胚胎上的毒物进行定量评估。4)斑马鱼转基因技术可应用于斑马鱼等本土物种。这项建议的具体目的是：1.探索使用转基因斑马鱼模型来筛选几种超级基金化学物质，以检测GFP表达的差异。研究GFP阳性原始生殖细胞的行为及其与发育中生殖道的结合情况，并通过特定目的研究的杀虫剂处理，了解是否对鱼类的性别决定有生物学影响。III.为适当的转基因启动子开发双顺反子载体，允许配体诱导表达荧光素酶基因(用于定量测量)和GFP基因(用于信号的解剖定位)。IV.根据特定目标I和II的结果，开发具有最适当转基因的Fundulus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.