Development of Combinatorial Technology To Simultaneously Accomplish Detection and Purification of the Heavy Metal Contamination.

低水平、大范围的大气重金属污染将严重危害人类健康，是未来的环境问题之一。在这项研究中，组合技术，同时检测和净化的重金属污染的发展。首先，通过在细菌细胞中引入重金属转运活性和报告基因表达系统，建立了重金属污染的灵敏检测系统。第二，将汞离子转运蛋白基因和多磷酸盐合成基因ppk基因导入烟草细胞，培育出具有重金属去除活性的植物。第三、为了了解汞离子转运蛋白基因在酵母细胞中的表达分布，研究了将汞离子转运蛋白基因导入酵母细胞的具体方法，并对所研制的汞离子生物传感器进行了实验研究，结果表明，该传感器具有很高的灵敏度（无机汞离子检测限为1 nM，有机汞化合物检测限为0.5 nM）并且GFP融合生物传感器装置不需要任何用于汞检测的特殊设备，并且其可适用于测量实际环境场所中的汞污染。从使用转基因植物（烟草）的研究中可以理解，带有汞转运蛋白基因merT的ppk基因克隆对于培育抗汞和积累汞的植物是有效的。利用酵母细胞表达系统的研究表明，所表达的重金属转运蛋白MerC与突触融合蛋白Vam3p的融合蛋白特异性地定位于酵母细胞的液泡膜上，赋予酵母细胞对重金属的抗性和积累能力。

As one of environmental problems in future, low level and wide aerial contamination of heavy metals will be serious for human health. In this study, combinatorial technology for simultaneous detection and purification of the heavy metal contamination was developed. First, sensitive detection system of contaminated heavy metals was developed by introducing heavy metal transportation activity and a reporter gene expression systems into bacterial cells. Second, plants that have heavy metal removal activity were developed using mercury ion transporter gene and polyphosphate synthesis gene, ppk gene, into tobacco cells. Third, specific methods of introduction of a mercury ion transporter gene into yeast cells were investigated to know maldistribution of the gene expression in the yeast cells.The experimental results obtained from the research showed that the developed mercury biosensor has very high sensitivity (with detection limit of 1 nM for inorganic mercury ion and 0.5 nM for organic mercurial compound) and that the gfp fusion biosensor device does not required any special equipment for the mercury detection and it can be applicable for measurement of the mercury contamination in actual environmental sites. From the study using transgenic plant (tobacco), it was understood that ppk gene cloning with a mercury transporter gene, merT, is effective to bread mercury resistant and accumulation plants. Form the study using yeast cell expression system, it was shown that the expressed fusion protein of heavy metal transporter protein, MerC, and a syntaxine family protein, Vam3p is specially located at the vacuole membrane of the yeast cell, and it confers heavy metal resistance and accumulation capability to the yeast cell.