Willardson B M, Wilkins J F, Rand T A, Schupp J M, Hill K K, Keim P, Jackson P J
Department of Chemistry and Biochemistry, Brigham Young University, Provo, Utah 84602, USA.
Appl Environ Microbiol. 1998 Mar;64(3):1006-12. doi: 10.1128/AEM.64.3.1006-1012.1998.
A bacterial biosensor for benzene, toluene, and similar compounds has been constructed, characterized, and field tested on contaminated water and soil. The biosensor is based on a plasmid incorporating the transcriptional activator xylR from the TOL plasmid of Pseudomonas putida mt-2. The XylR protein binds a subset of toluene-like compounds and activates transcription at its promoter, Pu. A reporter plasmid was constructed by placing the luc gene for firefly luciferase under the control of XylR and Pu. When Escherichia coli cells were transformed with this plasmid vector, luminescence from the cells was induced in the presence of benzene, toluene, xylenes, and similar molecules. Accurate concentration dependencies of luminescence were obtained and exhibited K1/2 values ranging from 39.0 +/- 3.8 microM for 3-xylene to 2,690 +/- 160 microM for 3-methylbenzylalcohol (means +/- standard deviations). The luminescence response was specific for only toluene-like molecules that bind to and activate XylR. The biosensor cells were field tested on deep aquifer water, for which contaminant levels were known, and were able to accurately detect toluene derivative contamination in this water. The biosensor cells were also shown to detect BETX (benzene, toluene, and xylene) contamination in soil samples. These results demonstrate the capability of such a bacterial biosensor to accurately measure environmental contaminants and suggest a potential for its inexpensive application in field-ready assays.
一种用于检测苯、甲苯及类似化合物的细菌生物传感器已构建完成、进行了表征，并在受污染的水和土壤中进行了现场测试。该生物传感器基于一种质粒，该质粒整合了来自恶臭假单胞菌mt-2的TOL质粒的转录激活因子xylR。XylR蛋白可结合一类甲苯样化合物，并在其启动子Pu处激活转录。通过将萤火虫荧光素酶的luc基因置于XylR和Pu的控制之下，构建了一个报告质粒。当用这种质粒载体转化大肠杆菌细胞时，在苯、甲苯、二甲苯及类似分子存在的情况下，细胞会发出荧光。获得了准确的荧光浓度依赖性，其K1/2值范围从3-二甲苯的39.0±3.8微摩尔到3-甲基苄醇的2,690±160微摩尔（平均值±标准差）。荧光响应仅对能结合并激活XylR的甲苯样分子具有特异性。该生物传感器细胞在已知污染物水平的深层含水层水中进行了现场测试，能够准确检测该水中的甲苯衍生物污染。该生物传感器细胞还被证明能够检测土壤样品中的BETX（苯、甲苯和二甲苯）污染。这些结果证明了这种细菌生物传感器准确测量环境污染物的能力，并表明其在现场即用型检测中具有廉价应用的潜力。
