Pérez-de-Mora Alfredo, Schulz Stephan, Schloter Michael
Department for Terrestrial Ecogenetics, Helmholtz Zentrum München, Institute of Soil Ecology, Neuherberg, Germany.
Methods Mol Biol. 2010;599:59-68. doi: 10.1007/978-1-60761-439-5_4.
Hydrocarbons are major contaminants of soil ecosystems as a result of uncontrolled oil spills and wastes disposal into the environment. Ecological risk assessment and remediation of affected sites is often constrained due to lack of suitable prognostic and diagnostic tools that provide information of abiotic-biotic interactions occurring between contaminants and biological targets. Therefore, the identification and quantification of genes involved in the degradation of hydrocarbons may play a crucial role for evaluating the natural attenuation potential of contaminated sites and the development of successful bioremediation strategies. Besides other gene clusters, the alk operon has been identified as a major player for alkane degradation in different soils. An oxygenase gene (alkB) codes for the initial step of the degradation of aliphatic alkanes under aerobic conditions. In this work, we present an MPN- and a real-time PCR method for the quantification of the bacterial gene alkB (coding for rubredoxin-dependent alkane monooxygenase) in environmental samples. Both approaches enable a rapid culture-independent screening of the alkB gene in the environment, which can be used to assess the intrinsic natural attenuation potential of a site or to follow up the on-going progress of bioremediation assays.
由于石油泄漏失控以及向环境中处置废物，碳氢化合物成为土壤生态系统的主要污染物。由于缺乏合适的预后和诊断工具来提供污染物与生物靶点之间发生的非生物 - 生物相互作用的信息，受影响场地的生态风险评估和修复常常受到限制。因此，鉴定和定量参与碳氢化合物降解的基因对于评估污染场地的自然衰减潜力以及制定成功的生物修复策略可能起着关键作用。除了其他基因簇外，alk操纵子已被确定为不同土壤中烷烃降解的主要参与者。一种加氧酶基因（alkB）编码有氧条件下脂肪族烷烃降解的第一步。在这项工作中，我们提出了一种MPN法和一种实时PCR法，用于定量环境样品中的细菌基因alkB（编码依赖于红氧还蛋白的烷烃单加氧酶）。这两种方法都能够对环境中的alkB基因进行快速的非培养筛选，可用于评估场地的内在自然衰减潜力或跟踪生物修复试验的进展情况。
