油藏环境中石油烃厌氧生物降解初始活化功能菌组成与活性

Anaerobic biodegradation of petroleum hydrocarbon is an important part of the natural carbon cycling, and the technology for oil extraction based on anaerobic biodegradation of residual oil and subsequent methane-production in oil reservoir plays a critical role in the deep exploitation of oilfields and sustainable development of our country. Initial activation of petroleum hydrocarbon is the key step for its anaerobic biodegradation. Focusing on the basic scientific issues encountered in anaerobic biodegradation of petroleum hydrocarbon, this study will analyze functional microorganisms involved in the initial activation of petroleum hydrocarbon and their abundance within microbial communities inhabiting oil reservoirs by analyzing related functional genes through designed PCR probes and q-PCR technology. The active microorganisms conducting initial activation of oil as well as their activity will be further analyzed by establishment of cDNA cloning libraries using reverse transcription of mRNA, together with RT-PCR and quantitative analysis of characteristic metabolites and etc. With the analytical results of the composition, abundance and gene transcripts of functional microorganisms involved in anaerobic biodegradation obtained from petroleum reservoirs with different temperature, salinity and biodegradation levels of crude oil, the relationship between the composition, abundance as well as the activity of these functional microorganisms and oil reservoir environmental conditions will be revealed by methods used for analysis of ecology diversity. Consequently, the comprehensive and systematic data will be obtained towards the functional microorganisms involved in the initial activation of anaerobic biodegradation of petroleum hydrocarbon. The results obtained will provide theoretical support on the study of biochemical mechanisms, thermodynamics and kinetics of anaerobic degradation of petroleum hydrocarbon. This research will lay a theoretical foundation for the development and utilization of microbial resources in oil reservoir, especially for the development and application of oil extraction technologies based on the biological gasification of residual oil in oil reservoirs.

石油烃厌氧生物降解是自然界碳循环的重要组成部分，以此为基础的残余油生物气化开采技术对促进我国石油资源深度开发和可持续发展具有重要意义。石油烃初始活化是其厌氧生物降解的前提和关键。本项目针对油藏中石油烃厌氧生物降解，优化相关功能基因探针，结合q-PCR，分析初始活化功能菌组成及其丰度；利用mRNA反转录cDNA克隆建库和RT-PCR技术，结合标志性代谢产物检测等，进一步鉴定分析油藏环境中实际行使初始活化功能的微生物及其活性；通过不同代表性油藏中初始活化功能菌组成、丰度及功能基因表达活性分析，借助分子生态学多样性分析原理方法，探索解析油藏中初始活化功能菌组成及其活性与油藏环境的关系。由此，获得油藏环境中行使石油烃厌氧生物降解初始活化功能微生物的系统的基础数据，为石油烃厌氧降解机制、热力学及动力学规律研究提供理论支持，为油藏微生物资源的高效利用及残余油生物气化开采技术开发奠定基础。

石油烃初始活化是石油烃厌氧生物降解过程的前提、核心和关键。解析并深入认识地下油藏中石油烃厌氧生物降解初始活化功能菌组成及其活性，对分析石油烃生物降解机制，研究地下环境中的碳循环生化反应过程具有重要的科学价值，对构建油藏残余油生物气化开采技术，最终大幅度提高石油资源的开发利用率具有重要的现实意义。.油藏环境中发育着种类繁多、功能多样的微生物，形成了相互关联、相互影响的生态系统，进行着多样的生物地球化学反应。本项目针对油藏石油烃厌氧降解，采集代表性油藏样品，采用assA及ahyA功能基因分析了烃厌氧降解起始活化功能菌；采用RT-PCR、cDNA克隆建库技术结合标志性代谢产物检测等分析明确了实际行使烃降解初始活化功能的微生物及其功能基因表达活性，在DNA、RNA水平及相应代谢产物三个方面获得了油藏中进行的石油烃厌氧初始活化过程的系统的、一致性的科学数据，充分地证实了油藏环境中活跃进行着烃厌氧降解初始活化及产甲烷过程，并分析归纳了其生化反应途径；以油藏产出液为基质，成功构建了烃厌氧降解生化反应模拟体系，采用DNA-SIP结合16S rRNA克隆建库技术，识别并鉴定了模拟体系中活跃参与正构十六烷降解的主要功能菌。本研究在油藏环境中石油烃厌氧生物降解初始活化菌群、生化反应过程及动力学方面方面取得了系统深入的科学数据，为相关研究及应用奠定了基础。

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