共查询到17条相似文献,搜索用时 82 毫秒
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研究确定土壤微生物基因组DNA提取方法、PCR扩增条件、DGGE电泳条件,为进一步研究分析土壤中微生物结构变化规律提供理论依据。土壤微生物基因组DNA提取采用直接法和间接法进行比较; PCR扩增条件调整扩增体系、DGGE电泳条件调整变性剂范围,并对其结果进行比较分析。通过对DGGE电泳相关条件的研究,结果显示,土壤中粗基因组DNA采用直接法提取,然后进行纯化; PCR扩增体系中加入BSA,DGGE电泳系统组成中变性剂浓度范围为35%~55%。确定了土壤微生物基因组DNA提取方法、PCR扩增条件、DGGE电泳条件,为后续的相关研究提供理论依据。 相似文献
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PCR-DGGE技术在农田土壤微生物多样性研究中的应用 总被引:43,自引:6,他引:43
变性梯度凝胶电泳技术(DGGE)在微生物生态学领域有着广泛的应用。研究采用化学裂解法直接提取出不同农田土壤微生物基因组DNA,并以此基因组DNA为模板,选择特异性引物F357GC和R515对16S rRNA基因的V3区进行扩增,长约230bp的PCR产物经变性梯度凝胶电泳(DGGE)进行分离后,得到不同数目且分离效果较好的电泳条带。结果说明,DGGE能够对土壤样品中的不同微生物的16S rRNA基因的V3区的DNA扩增片断进行分离,为这些DNA片断的定性和鉴定提供了条件。与传统的平板培养方法相比,变性梯度凝胶电泳(DGGE)技术能够更精确的反映出土壤微生物多样性,它是一种有效的微生物多样性研究技术。 相似文献
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应用变性梯度凝胶电泳和16SrDNA序列分析对kefir粒中细菌多样性的研究 总被引:2,自引:0,他引:2
以开菲尔(Kefir)粒为材料,经过DNA抽提和16SrDNA V3区PCR扩增,扩增产物经变性梯度凝胶电泳(DGGE)分离并切割电泳条带进行序列测定,并与现有的数据库进行了比较,对Kefir粒的细菌多样性进行分析。结果表明,DGGE图谱中可检测到的8条带的16SrDNA基因序列中有7个基因序列与GenBank数据库登录的相关序列的相似性大于98%,余下的1个基因序列的相似性也大于96%。相似性大于98%的7个克隆中,有3个属于鞘氨醇杆菌属(Sphingobacterium),2个属于乳杆菌属(Lactobacillus),其它2个分别属于肠杆菌属(Errterobacter)和不动杆菌属(Acinetobacter)。首次报道了鞘氨醇杆菌作为优势菌群存在开菲尔Kefir粒中。 相似文献
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变性梯度凝胶电泳在微生物分子生态学研究中的应用日益广泛,已成为研究微生物多样性和动态变化的有力工具.对变性梯度凝胶电泳技术中存在的问题,如基因片段的选择、共迁移、背景色和异源双链分子的形成等作了综述,以期为进一步更好地利用该技术进行微生态的研究奠定基础. 相似文献
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变性梯度凝胶电泳在环境微生物生态学中的应用 总被引:1,自引:2,他引:1
PCR-变性梯度凝胶电泳(PCR-DGGE)具有可靠性强、重复性好、方便快捷等优点,已被广泛应用于环境生态学中微生物群落多样性、动态性分析和功能细菌的跟踪。本文综述了PCR-DGGE技术的基本原理,不同DNA提取方法的比较,不同PCR方式的比较及其在环境生态学中研究微生物群落多样性、环境中微生物群落变化的动态监测、硝化菌-反硝化菌和硫酸还原菌(SRB)的动态分析和监测等领域中的应用,并对该技术自身存在的局限性和应用前景进行了评价。 相似文献
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目的 研究变性梯度凝胶电泳(denatured gradient gel electrophoresis, DGGE)在实验小鼠细菌检测中的应用。方法 根据16S rDNA V3区引物,PCR扩增3种实验小鼠(KM小鼠、NIH小鼠和BALB/c小鼠)呼吸道和盲肠段的细菌基因组DNA;扩增产物运用DGGE进行电泳检测,并分析条带数量间差异的统计学意义。结果 KM小鼠盲肠段条带12~18条,呼吸道条带5~10条;NIH小鼠盲肠段条带15~20条,呼吸道条带4~10条;BALB/c小鼠盲肠段条带10~15条,呼吸道条带0~7条。统计分析结果显示,KM小鼠和NIH小鼠在盲肠和呼吸道电泳条带数量上的差异无统计学意义( P >0.05);BALB/c小鼠与KM小鼠、NIH小鼠间的差异均有统计学意义( P <0.05)。结论 DGGE 在实验小鼠盲肠和呼吸道细菌检测中能较好地反映菌群的物种多样性。 相似文献
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DGGE/TGGE技术及其在微生物分子生态学中的应用 总被引:48,自引:1,他引:48
变性梯度凝胶电泳(DGGE)和温度梯度凝胶电泳(TGGE)是近些年微生物分子生态学研究中的热点技术之一。由于DGGE/TGGE技术具有可靠性强、重现性高、方便快捷等优点,被广泛地应用于微生物群落多样性和动态性分析。文章对DGGE/TGGE技术原理与关键环节、局限性和应用前景进行了综述。 相似文献
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Microbial communities within subsurface uranium (U) deposits were explored to understand the nature of community composition and their potential role in biogeochemical cycle and bioremediation. Geochemical analysis revealed that the U ores were mainly hosted on metamorphosed chlorite-biotite schists, containing varied organic carbon and elevated level of several heavy metals (U, Cu, Cr, Zn, etc.). Microbial diversity as explored by 16S rRNA gene clone library and DGGE analyses revealed predominance of Proteobacteria, Acidobacteria, Bacteroidetes along with Firmicutes and candidate division OP9 within the domain Bacteria and Euryarchaeota within the domain Archaea. Among the physiochemical parameters, level of organic carbon showed considerable impact on influencing community diversity and composition. Samples from Jaduguda with high organic carbon showed abundance of bacteria known for metabolizing different carbon compounds and affiliated to unclassified uncultured members of Chitnophagaceae (Bacteroidetes), Gp4 of Acidobacteria and unclassified β-Proteobacteria and halophilic, nitrate-reducing γ-Proteobacteria. A relatively diverse assemblage of species capable of autotrophic/heterotrophic N2 fixation, CH4 utilization, H2(0)/Fe(II)/Mn(II)/S(0) oxidation, NO3/Fe(III) reduction, U and other metal precipitation/mineralization and affiliated to families Rhizobiaceae, Bradyrhizobiaceae, Caulobacteraceae, Comamonadaceae and genera Acinetobacter, Marinobacter and Alcanivorax constituted the Bagjata samples. Distribution and interrelations among abundance of various bacterial groups detected in other U mines/radioactive waste sites were compared with our data. Overall, the study reported distinct compositions of indigenous microbial communities among the samples from two mines and provided a better insight in geomicrobiology of U deposits. 相似文献
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微生物物种多样性的保护与其资源保藏 总被引:13,自引:0,他引:13
从微生物物种多样性的保护以及微生物物种资源的保藏等方面较全面地阐述了微生物物种资源保护与保藏的重要性及其相互关系 ,阐明了微生物菌种保藏的原理及方法 ,同时介绍了保藏技术领域的研究动向和保藏机构或组织在微生物资源的保藏、开发和交流等方面所起的重要作用 相似文献
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Nested PCR-Denaturing Gradient Gel Electrophoresis Approach To Determine the Diversity of Sulfate-Reducing Bacteria in Complex Microbial Communities 总被引:4,自引:0,他引:4 下载免费PDF全文
Here, we describe a three-step nested-PCR-denaturing gradient gel electrophoresis (DGGE) strategy to detect sulfate-reducing bacteria (SRB) in complex microbial communities from industrial bioreactors. In the first step, the nearly complete 16S rRNA gene was amplified using bacterial primers. Subsequently, this product was used as a template in a second PCR with group-specific SRB primers. A third round of amplification was conducted to obtain fragments suitable for DGGE. The largest number of bands was observed in DGGE patterns of products obtained with primers specific for the Desulfovibrio-Desulfomicrobium group, indicating a large diversity of these SRBs. In addition, members of other phylogenetic SRB groups, i.e., Desulfotomaculum, Desulfobulbus, and Desulfococcus-Desulfonema-Desulfosarcina, were detected. Bands corresponding to Desulfobacterium and Desulfobacter were not detected in the bioreactor samples. Comparative sequence analysis of excised DGGE bands revealed the identity of the community members. The developed three-step PCR-DGGE strategy is a welcome tool for studying the diversity of sulfate-reducing bacteria. 相似文献
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为了强调物种多样性在微生物学课程中的分量,近几年我们尝试对教学内容进行了改革:①将微生物分类单元、细胞生物的拉丁双名法等共性分类知识调到绪论中;②在讲授原核微生物、真核微生物和病毒各章时,分别增加了它们的多样性与分类内容;③考虑到课时所限,采用图表概括方式介绍各大类微生物的主要分类类群及主要特征,将原先课堂教学中介绍的一些特殊微生物内容纳入到微生物多样性体系中讲解,适当予以详细介绍, 相似文献
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GC Fractionation Enhances Microbial Community Diversity Assessment and Detection of Minority Populations of Bacteria by Denaturing Gradient Gel Electrophoresis 总被引:1,自引:3,他引:1 下载免费PDF全文
William E. Holben Kevin P. Feris Anu Kettunen Juha H. A. Apajalahti 《Applied microbiology》2004,70(4):2263-2270
Effectively and accurately assessing total microbial community diversity is one of the primary challenges in modern microbial ecology. This is particularly true with regard to the detection and characterization of unculturable populations and those present only in low abundance. We report a novel strategy, GC fractionation combined with denaturing gradient gel electrophoresis (GC-DGGE), which combines mechanistically different community analysis approaches to enhance assessment of microbial community diversity and detection of minority populations of microbes. This approach employs GC fractionation as an initial step to reduce the complexity of the community in each fraction. This reduced complexity facilitates subsequent detection of diversity in individual fractions. DGGE analysis of individual fractions revealed bands that were undetected or only poorly represented when total bacterial community DNA was analyzed. Also, directed cloning and sequencing of individual bands from DGGE lanes corresponding to individual G+C fractions allowed detection of numerous phylotypes that were not recovered using a traditional random cloning and sequencing approach. 相似文献