| 基于荧光传感器Frex的特性检测水质环境中生物毒性物质 |
| 摘要点击 2465 全文点击 1499 投稿时间：2013-10-11 修订日期：2013-12-13 |
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| 中文关键词 Frex 荧光发光菌 毒物测试 水质环境 条件优化 |
| 英文关键词 Frex luminescent bacteria toxicity test water environment condition optimization |
| 作者 | 单位 | E-mail | | 赵巍 | 浙江工业大学生物与环境工程学院, 杭州 310014
浙江清华长三角研究院, 浙江省应用酶学重点实验室, 嘉兴 314006 | zw163163@yeah.net | | 汪钊 | 浙江工业大学生物与环境工程学院, 杭州 310014 | | | 蔡强 | 浙江清华长三角研究院, 浙江省应用酶学重点实验室, 嘉兴 314006 | | | 欧文斌 | 浙江清华长三角研究院, 浙江省应用酶学重点实验室, 嘉兴 314006 | | | 孟凡国 | 浙江清华长三角研究院, 浙江省应用酶学重点实验室, 嘉兴 314006 | mengfanguo@tsinghua.org.cn |
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| 中文摘要 |
| 发光菌作为水环境生物毒性监控的有效手段，近年来越来越得到重视. 本实验利用基因工程技术，将Frex荧光蛋白重组到大肠杆菌中，并通过测定菌体内NADH水平所引起的荧光强度变化，来监测水体中有毒物质对微生物代谢的影响，从而实现实时毒性监控. 实验完成了Frex荧光蛋白在大肠杆菌中的正确表达，研究了培养温度、诱导时间及IPTG浓度等不同条件对Frex融合蛋白表达量和荧光活性的影响. 将此重组荧光发光菌初步应用于环境水体中有毒物质的检测研究，选取氯化汞、苯酚、重铬酸钾、硫酸锌等4种国际荧光发光菌生物毒性检测标准物质进行检测，结果表明毒性物质均能对菌体的荧光产生一定的催灭作用. 该毒物测试手段分别具有反应速度快和灵敏度高等特性. 同时，实验优化了该重组菌应用于生物毒性检测试验的条件，为后续的应用研究提供了一定的实验基础，拓展了荧光发光菌在其他方面的适用范围. |
| 英文摘要 |
| Luminescent bacteria have attracted more and more attention in recent years as an effective mean for biological toxicity of water environment monitoring. First of all, fluorescent protein Frex was correctly expressed in Escherichia coli, and then the effect of toxic substances on microbial metabolism in the water was monitored through the determination of the changes in the fluorescence intensity in bacteria caused by the change of NADH level in the bacteria. Then the effects of culture temperature, inducing time and the final concentration of inductor isopropyl beta-D-thiogalactopyranoside (IPTG) on the expression level and fluorescent activity of the fusion protein Frex were studied. The recombinant fluorescent bacteria was then applied in the initial detection of toxic substances in water environment. Four international standard substances of biological toxicity test including HgCl2, 3,5-dichlorophenol, potassium dichromate, and zinc sulfate heptahydrate were chosen to conduct experimental assay. The results suggested that all of these substances can cause a rapid decrease in the fluorescence of the bacteria. This test method has advantages of rapid reaction and high sensitivity. Meanwhile, the optimization of the conditions for the biological toxicity test lays foundation for subsequent application, and expands the application scope of luminescent bacteria in other aspects. |
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