Division of Applied Life Science and Institute of Agriculture and Life Sciences, Gyeongsang National University, Jinju 660-701, Republic of Korea.
Biosens Bioelectron. 2013 Dec 15;50:256-61. doi: 10.1016/j.bios.2013.06.058. Epub 2013 Jul 4.
In this study, we developed a simple and sensitive biosensor for the determination of toxoflavin (which is toxic to various plants, fungi, animals, and bacteria) in natural samples based on β-galactosidase activity. The proposed toxoflavin detection method for toxin-producing bacteria or toxin-contaminated foods is simple and cost effective. Burkholderia glumae, a species known to cause rice grain rot and wilt in various field crops, produces toxoflavin under the control of a LysR-type transcriptional regulator ToxR and its ligand toxoflavin. As the expression of toxoflavin biosynthetic genes requires toxoflavin as a co-activator of ToxR, a novel biosensor stain was constructed based on lacZ reporter gene integration into the first gene of the toxoflavin biosynthesis operon, toxABCDE of B. glumae. The biosensor was composed of a sensor strain (COK71), substrates (X-gal or ONPG), and culture medium, without any complex preparation process. We demonstrated that the biosensor strain is highly specific to toxoflavin, and can quantify relative amounts of toxoflavin compared with known concentrations of toxoflavin. The proposed method was reliable and simple; samples containing 50-500 nM of toxoflavin could be analyzed. More importantly, the proposed biosensor strain could identify toxoflavin-producing bacteria in real samples. The excellent performance of this biosensor is useful for diagnostic purposes, such as detecting toxoflavin-contaminated foods and environmental samples.
在本研究中，我们基于β-半乳糖苷酶活性开发了一种用于测定曲酸（对各种植物、真菌、动物和细菌均有毒性）的简单、灵敏的生物传感器，用于天然样品中曲酸的测定。本研究提出的产毒细菌或毒素污染食品的曲酸检测方法简单且经济有效。能引起水稻粒腐病和多种田间作物萎蔫的恶臭假单胞菌（Burkholderia glumae）在 LysR 型转录调控因子 ToxR 及其配体曲酸的控制下产生曲酸。由于曲酸生物合成基因的表达需要曲酸作为 ToxR 的共激活剂，因此根据恶臭假单胞菌 toxABCDE 基因簇的第一个基因构建了新型的曲酸生物合成基因生物传感器。生物传感器由一个传感器菌株（COK71）、底物（X-gal 或 ONPG）和培养基组成，无需任何复杂的制备过程。结果表明，生物传感器菌株对曲酸具有高度特异性，与已知浓度的曲酸相比，能够定量相对量的曲酸。该方法可靠且简单，能够分析含有 50-500 nM 曲酸的样品。更重要的是，该生物传感器菌株能够在实际样品中识别产曲酸细菌。该生物传感器具有良好的性能，可用于诊断目的，如检测曲酸污染食品和环境样品。
