荧光原位杂交技术及其在微生物生态学中的应用  

Fluorescence in situ hybridization (FISH) and its applications in microbial ecology

作　　者：呼庆[1] 齐鸿雁[1] 张洪勋[1] 

机构地区：[1]中国科学院生态环境研究中心环境生物技术研究室,北京100085

出　　处：《生态学报》2004年第5期1048-1054,共7页Acta Ecologica Sinica

基　　金：国家"十五"科技攻关重点资助项目 ( 2 0 0 1BA90 3 B)~~

摘　　要：综述了荧光原位杂交技术 (fluorescence in situ hybridization FISH)在微生物生态学领域的各种应用 ,同时就其发展过程。During recent years,molecular techniques such as PCR and denaturing gradient gel electrophoresis (DGGE) or DNA sequencing have revolutionized all fields of microbiology,and sensitive detection and exact identification of bacteria are possible.Fluorescence in situ hybridization (FISH) using 16SrRNA probes does not rely on PCR amplification,and as such provides a useful complementary technique to DGGE for the analysis of organisms. Because of FISH allowing nucleic acid sequences to be examined inside cells without altering the cell's morphology or the integrity of its various compartments and providing information about number,spatial distribution and cellular environment,it has become a powerful tool for phylogenetic,ecologic,diagnostic and environmental studies in microbiology. Fluorescence in situ hybridization can detect nucleic acid sequences by a fluorescently labeled probe that hybridizes specifically to its complementary target sequence within the intact cell. Its general procedure in FISH analysis of microorganisms is as follows: (1) fixation of the specimen; (2) preparation of the sample,possibly including specific pretreatment steps; (3) hybridization with the respective probes for detecting the respective target sequences; (4) washing steps to remove unbound probes; (5) mounting,visualization and documentation of results. Because FISH gives a detailed picture of the microenviroments without any selective purification or amplification steps, there is a large scope of FISH applications. It has been extensively used in the field of environmental microorganisms diversity such as the investigation of microbial communities of aquatic habitats and soil habitats. Most obligate microbial symbionts are as-yet uncultured. Using the 16SrRNA approach,they can be identified and phylogenetically classified.Using different FISH strategies,bacterial endosymbionts were detected in many microorganisms. Population analysis by FISH has proven particularly useful for description of the normal flora or that of mixed mic

关 键 词：荧光原位杂交 微生物生态 16SrRNA 探针 

分 类 号：Q93-3[生物学—微生物学] Q938