Research on the new biotechnology for specific detection of environmental contamination by hazardous heavy metals.

土壤和水体的重金属污染是世纪以来工业化国家普遍面临的严重环境问题。最近，这一问题被认为是发展中国家非常危险的健康问题。为了预防和解决这一环境问题，我们的目标是开发生物传感器技术，利用细菌细胞诱导的基因表达系统选择性检测影响健康的重金属。本研究的目的是开发比传统分析方法更灵敏的重金属生物传感器。(1)以细菌发光基因（lux）为报告基因，建立了一种环境中有机汞化合物的检测系统：利用广谱抗汞细菌巨大芽孢杆菌（Bacillusmagaterlum）MB 1的一种新的有机汞裂解酶基因merB's，构建了merB 3-荧光素酶（mer-lux）融合转录质粒。另一个质粒编码来自相同B的操纵子/启动子序列和mer操纵子基因。megaterlum MB 1的构建，并用作在土方Ichla科尔细胞中与mer-lux融合质粒的反式作用基因表达载体。含不同merB基因融合质粒和lux基因表达质粒的大肠杆菌对烷基汞和芳香族汞的反应不同。(2)利用新研制的生物传感器装置，可以检测到极低浓度的无机汞。用lux基因、merR和merTgenes基因表达质粒转化的细菌细胞显示pM水平的汞污染。该灵敏度比原子吸收法、离子耦合等离子体质谱法等以往的分析方法高约100倍。

Heavy metal contamination of soil or water has been frequent as a severe environmental problem in industrialized countries through th 20 century. Recently, this problem is recognized as a very dangerous health problem in developing countries. To prevent and solve this environmental problem, we aimed development of biosensor technology for selective detection of health affecting heavy metals using an induced gene expression system by bacterial cells. This research work was done to develop more sensitive heavy metal biosensors than conventional analytical methods.The two major results derived from this research project are as follows.(1) Using bacterial luminescence genes (lux) as reporter genes, we developed a detection system of organomercurial compounds in the environment: A new organomercury lyase genes, merB's, from a broad-spectrum mercury resistant bacterial strain, Bacillus magaterlum MB1, was used to construct merB3-luciferase (mer-lux) transcriptional fusion plasmids. Another piasmid encoding an operator/promoter sequence and mer operon genes from the same B. megaterlum MB1 was constructed and used as a transacting gene expression vector with the mer-lux fusion plasmids in a Escherlchla coll cell. The transformant with different merB genes fusion plasmids and a lux gene expression plasmids showed deferent response to alkyl-or aromatic-organomercurials tested in this study.(2) Very low concentration of inorganic mercury could be detected by newly developed biosensor devise bacterial cells. The bacterial cells transformed with lux genes, merR and merTgenes gene expression plasmids showed pM level mercury contamination. This sensitivity is about one hundred times higher than those of conventional analytical methods such as an atomic absorption method or an ion coupling plasma-mass spectrometory.