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基于PCR-DGGE基因指纹的对虾体内优势细菌组成分析 总被引:16,自引:0,他引:16
采用不依赖分离培养的16S rDNA的PCR-DGGE基因指纹技术对刀额新对虾与中国对虾的鳃部与肠道优势细菌种群组成进行比较分析。研究发现:对虾鳃部与肠道存在着丰富多样的细菌;根据DGGE指纹图的聚类分析发现不同对虾及同一种对虾的鳃部与肠道内的细菌组成差异性非常大;同时也发现不同对虾体内有相同的细菌存在。首次尝试建立基于16S rDNA的PCR-DGGE基因指纹的对虾体内细菌组成揭示方法,对于今后建立对虾与养殖水体微生物和相关疾病的关系具有重要意义。 相似文献
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Sun YZ Mao SY Yao W Zhu WY 《Animal : an international journal of animal bioscience》2008,2(3):391-398
In the rumen, plant particles are colonised and degraded by the rumen micro-organisms. Although numerous important findings about fibre-associated bacterial community were obtained using traditional or molecular techniques, little information is available on the dynamics of bacteria associated with feed particles during incubation in the rumen. In the present study, ryegrass leaf, ryegrass stem and rice straw, representing different carbohydrate compositions, were used as substrates and placed in the rumen of goats by using nylon bags, and PCR/DGGE (denaturing gradient gel electrophoresis) with subsequent sequence analysis were used to monitor the dynamics of and identify bacteria associated with the substrates during 24 h of incubation. DGGE results showed that substrate samples collected from 10 min to 6 h had similar DGGE patterns, with up to 24 predominant bands to each sample, including 14 common bands to all samples, suggesting a rapid and stable colonisation by a highly diverse bacterial community. Substrate samples collected at 12 and 24 h showed similar DGGE patterns but had great difference in DGGE patterns from those collected at 10 min to 6 h, suggesting an apparent shift in bacterial community. Sequence analysis indicated that most substrate-associated bacteria were closely related to fibrolytic bacteria. In conclusion, a highly diverse and similar rumen bacterial community could immediately colonise to different substrates and remained stable during the initial 6 h of incubation, but experienced a marked change after 12 h of incubation. Italian ryegrass leaf, Italian ryegrass stem and rice straw were colonised with a similar bacterial community. 相似文献
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We describe a simple method, based on heteroduplex mobility analysis of 16S rDNA fragments, for targeted detection of sub-populations of bacteria within diverse microbial communities. A small (ca. 200 bp) polymorphic fragment of the bacterial 16S rRNA gene was amplified from sample DNA using universal primers. Sample products were hybridised with a fluorescently labelled fragment amplified from a selected 'reporter' organism representing the target group. The resulting products were resolved and the labelled heteroduplex pairs detected on non-denaturing gels using automated DNA detection technology. A model, based on analysis of samples with known 16S rDNA sequences, demonstrates that heteroduplex mobility is inversely correlated with genetic distance and that beyond 26% genetic difference, heteroduplex products are not detected. The utility of the method was tested by field studies in which stream biofilms could be characterised by heteroduplex profiles generated with heterotrophic and autotrophic reporter organisms representing target groups. 相似文献
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Liu D Zhou Y Li C Li Y Jiang Y Huang Z Liang J Shu R 《Microbiology and immunology》2010,54(11):702-706
DGGE of 16S rDNA is one of the most frequently used methods to study microbial communities. In this study, the DGGE profiles of different 16S rDNA regions of the periodontal pathogens Porphyromonas gingivalis, Fusobacterium nucleatum, and Prevotella nigrescens were investigated. The results suggested that V3-V5 and V6-V8 fragments may be suitable for community analysis of subgingival bacteria. Further analysis of subgingival samples with V3-V5 and V6-V8 regions as target fragments suggested that, in chronic periodontitis, re-colonization by periodontal bacteria with a population very similar to the baseline may occur by 6 weeks after mechanical debridement. 相似文献
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Analysis of microbial diversity in oligotrophic microbial fuel cells using 16S rDNA sequences 总被引:14,自引:0,他引:14
Molecular ecological techniques were applied to analyze the bacterial diversity of two oligotrophic microbial fuel cells (MFCs) enriched using river water or artificial wastewater (AWW) as fuel. Denaturing gradient gel electrophoresis (DGGE) of the PCR amplified 16S rDNA showed that different microbial communities were present in the two MFCs and these were different from the river sediment used to initiate the enrichment. Nearly complete 16S rDNA was amplified and sequenced. Over 80% of the clones were Proteobacteria. Betaproteobacteria were the dominant clones (46.2%) in MFCs fed with river water, and about 64.4% of the clones in MFCs fed with AWW were Alphaproteobacteria. Actinobacteria were found only in the MFC fed with AWW, and Deltaproteobacteria, Acidobacteria, Chloroflexi and Verrucomicrobia in the MFC fed with river water. Many clones were related to uncultured bacteria, some with homology less than 95%, indicating that many novel bacteria were enriched in the oligotrophic MFCs. 相似文献
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茶园土壤微生物群落基因多样性 总被引:10,自引:0,他引:10
应用PCR技术,直接从土壤中抽提总DNA,扩增16S rDNA V3 片段,采用变性梯度凝胶电泳(DGGE)分析16S rDNA V3 片段的多态性,研究了杭州西湖梅家坞不同植茶年龄(8、50和90年)、不同利用方式(茶园、荒地和林地)的土壤微生物群落基因多样性.结果表明:不同植茶年龄和不同土地利用方式影响土壤微生物群落的基因多样性.荒地、茶园和林地土壤微生物群落基因多样性指数明显不同(P<0.05),其排列顺序为荒地>茶园>林地.不同植茶年龄的土壤中,50年茶园土壤的微生物群落基因多样性指数、微生物量碳和基础呼吸明显高于8年和90年茶园土壤(P<0.05). 相似文献
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Ji NN Liu MM Huang XR Zhen J Li SS Jiang S Yu HH Wang SY Peng XX 《Journal of applied microbiology》2006,100(4):795-799
AIMS: To develop an immunocapture universal primer PCR (iUPPCR) combined with denaturing gradient gel electrophoresis (DGGE) and evaluate it as a method permitting rapid detection of Shigella species and their serotypes. METHODS AND RESULTS: This method amplifying the conserved regions of bacterial 16S rRNA genes of different species or serotypes of Shigella dysentery bacilli captured and enriched by polyvalent antibodies can detect and distinguish causative pathogens rapidly. Four serotypes from three Shigella species including Shigella dysenteriae serotype 1, Shigella boydii serotype 1, Shigella flexneri serotypes 1a and 3a were examined. CONCLUSION: Our approach could be adopted for not only axenic bacterial population but also mixed communities and achieve rapid detection of various bacteria from the same genus or species in one sample. SIGNIFICANCE AND IMPACT OF THE STUDY: The iUPPCR-DGGE method was shown to be more convenient than serotype-specific-antibody-based method of iUPPCR for Shigella species detection and it could be also applied to the quick detection for other kinds of pathogens with many serotypes. 相似文献
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藏灵菇微生物种群结构的分子特性研究 总被引:7,自引:0,他引:7
用PCR—DGGE指纹技术,研究了藏灵菇中微生物多样性及藏灵菇发酵奶发酵过程微生物种群动力学。结果表明,藏灵菇中细菌的种群结构较酵母菌的复杂,不同来源的藏灵菇中细菌种群结构的相似性为78%-84%,酵母菌种群结构的相似性为80%-92%。发酵过程中细菌种群结构变化图谱中的条带B和条带E,以及酵母菌种群结构变化图谱中的条带N贯穿于整个发酵过程,是发酵过程的优势菌。序列分析表明,细菌种群结构的DGGE图谱中的绝大多数条带与乳酸菌相对应,其中最亮条带(条带E)的序列与乳酸乳球菌的相似性为100%。 相似文献
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Characterization of bacterial diversity in two aerated lagoons of a wastewater treatment plant using PCR-DGGE analysis 总被引:4,自引:0,他引:4
Moura A Tacão M Henriques I Dias J Ferreira P Correia A 《Microbiological research》2009,164(5):560-569
Aerated lagoons are commonly used for domestic and industrial wastewater treatment due to their low cost and minimal need of operational requirements. However, little information is known regarding microbial communities that inhabit these ecosystems. In this study, a 16S-DGGE approach was used to estimate bacterial diversity and to monitor community changes in two aerated lagoons from a wastewater treatment plant receiving urban and industrial effluents. Pronounced shifts between bacterial communities collected in winter–spring and summer–autumn months were detected. Temperature, dissolved oxygen (DO) and pH were the variables that most influenced the bacterial communities. Phylogenetic affiliation of predominant members was assessed by the determination of the 16S rDNA sequence of correspondent bands. Affiliations to Cytophaga–Flexibacter–Bacteroides (CFB) group, Firmicutes, and β- and ε-proteobacteria were found. 相似文献
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Jun Wang Ting Ma Lingxia Zhao Jinghua Lv Guoqiang Li Fenglai Liang Rulin Liu 《World journal of microbiology & biotechnology》2008,24(9):1981-1987
Different PCR–denaturing gradient gel electrophoresis (DGGE) protocols were employed to investigate bacterial communities
in a high temperature and water flooded petroleum reservoir in Dagang oil field, China. Bacterial universal primers sets frequently
used in PCR–DGGE were evaluated. Three primers sets P1 (341F-GC and 534R), P2 (341F-GC and 907R) and P3 (1055F and 1406R-GC)
showed different DGGE patterns. Good separation and quality of patterns were obtained in DGGE analysis with the set P3. A
total of 12 DNA fragments were excised from the DGGE gels and their sequences were determined. Clustering analysis of the
DGGE profiles showed that bacteria in this petroleum reservoir belonged to four clusters. These results indicate that the
procedure of DGGE analysis with the primer P3 (1055F and 1406R-GC) is suitable for investigating microbial community in petroleum
reservoirs. 相似文献
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Kyu-Jung Chae Mi-Jin ChoiJin-Wook Lee Kyoung-Yeol KimIn S. Kim 《Bioresource technology》2009,100(14):3518-3525
Four microbial fuel cells (MFCs) were inoculated with anaerobic sludge and fed four different substrates for over one year. The Coulombic efficiency (CE) and power output varied with different substrates, while the bacterial viability was similar. Acetate fed-MFC showed the highest CE (72.3%), followed by butyrate (43.0%), propionate (36.0%) and glucose (15.0%). Glucose resulted in the lowest CE because of its fermentable nature implying its consumption by diverse non-electricity-generating bacteria. 16S rDNA sequencing results indicated phylogenetic diversity in the communities of all anode biofilms, and there was no single dominant bacterial species. A relative abundance of β-Proteobacteria but an absence of γ-Proteobacteria was observed in all MFCs except for propionate-fed system in which Firmicutes dominating. The glucose-fed-MFC showed the widest community diversity, resulting in the rapid generation of current without lag time when different substrates were suddenly fed. Geobacter-like species with the most representative Geobactersulfurreducens PCAT were integral members of the bacterial community in all MFCs except for the propionate-fed system. 相似文献
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利用PCR-DGGE未培养技术对中国白酒高温和中温大曲细菌群落结构的分析 总被引:11,自引:2,他引:9
采用聚合酶链式反应-变性梯度凝胶电泳(PCR-DGGE)技术,研究了5种高温和中温白酒大曲细菌群落结构,通过优势条带切胶鉴定确定了大曲中优势细菌种属信息。结果表明,Weissella cibaria、Lactobacillus helveticus、L.fermentum、L.panis等乳酸菌普遍存在于5种大曲中,Ther-moactinomyces sanguinis仅存在于高温曲酱曲中,同时DGGE检测到了传统方法未能分离鉴定的Staphylococcus xylosus、Klebsiella oxytoca。不同工艺大曲细菌群落结构存在明显差异,随着制曲温度的升高,大曲细菌多样性指数有下降趋势。PCR-DGGE技术是一种能够快速有效地研究白酒大曲细菌群落结构的技术。 相似文献
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De Felice B Argenziano C Guida M Trifuoggi M Russo F Condorelli V Inglese M 《Molecular biotechnology》2009,41(2):123-132
Little is known about pharmaceutical and personal care products pollutants (PPCPs), but there is a growing interest in how they might impact the environment and microbial communities. The widespread use of Viagra (sildenafil citrate) has attracted great attention because of the high usage rate, the unpredictable disposal and the unknown potential effects on wildlife and the environment. Until now information regarding the impact of Viagra on microbial community in water environment has not been reported. In this research, for the first time, the genetic profile of the microbial community, developing in a Viagra polluted water environment, was evaluated by means of the 16S and 18S rRNA genes, for bacteria and fungi, respectively, amplified by polymerase chain reaction (PCR) and separated using the denaturing gradient gel electrophoresis (DGGE) technique. The DGGE results revealed a complex microbial community structure with most of the population persisting throughout the experimental period. DNA sequences from bands observed in the different denaturing gradient gel electrophoresis profiles exhibited the highest degree of identity to uncultured bacteria and fungi found previously mainly in polluted environmental and treating bioreactors. Biotransformation ability of sildenafil citrate by the microbial pool was studied and the capability of these microorganisms to detoxify a polluted water ecosystem was assessed. The bacterial and fungal population was able to degrade sildenafil citrate entirely. Additionally, assays conducted on Daphnia magna, algal growth inhibition assay and cell viability determination on HepG2 human cells showed that biotransformation products obtained from the bacterial growth was not toxic. The higher removal efficiency for sildenafil citrate and the lack of toxicity by the biotransformation products obtained showed that the microbial community identified here represented a composite population that might have biotechnological relevance to retrieve sildenafil citrate contaminated sites. 相似文献
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石油降解菌的分离鉴定及石油污染土壤的细菌多样性 总被引:19,自引:0,他引:19
从石油污染的土壤中分离筛选到28株石油降解菌,经鉴定分别为短杆菌属、假单胞菌属、邻单胞菌属和微球菌属;对4个石油不同程度污染的土壤样品中嗜油微生物分布状况进行分析,发现污染严重的土壤样品中嗜油菌的数量相对较多;用聚合酶链式反应(PCR)、变性梯度凝胶电泳(DGGE)和切胶测序相结合的方法对4个土壤样品中的细菌多样性进行分析,结果显示在受污染的土壤中,My cobacterium和B acillus在污染程度较低的样品中分布的较为集中,F lavobacterium和A zosp ira在污染程度较高的样品中丰度较高。属于B eta p roteobacterium类群的细菌在受污染的土壤中占有优势,同时还有一些不可培养的菌群存在。气质联用(GC-M S)分析结果表明石油污染程度及污染物中芳香烃类的含量对细菌多样性有着显著影响。在石油污染程度高,芳香烃类含量高的样品中细菌的多样性相对较低。 相似文献
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Johnny Kashama Véronique Prince Anne-Marie Simao-Beaunoir Carole Beaulieu 《Journal of industrial microbiology & biotechnology》2009,36(3):391-399
Forty-one bacterial strains isolated from the headbox water of two machines in a Canadian paper mill were associated with
the genera Asticcacaulis, Acidovorax, Bacillus, Exiguobacterium, Hydrogenophaga, Pseudomonas, Pseudoxanthomonas, Staphylococcus, Stenotrophomonas based on the sequence of their 16S rRNA genes. The metabolic profile of these strains were determined using Biolog EcoPlate,
and the bacteria were divided into four metabolic groups. Metabolic profiles of the bacterial communities colonizing the headbox
water of two paper machines was also determined weekly over a 1 year period. The only compound that was not reduced by the
bacterial community was 2-hydroxybenzoic acid. Utilization frequency of the other carbon sources in the Biolog EcoPlate ranged
from 3 to 100%. The metabolic profiles of the bacterial community did not vary considerably between the two paper machines.
However, the metabolic profile varied among the sampling dates. 相似文献
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DG-DGGE分析产氢发酵系统微生物群落动态及种群多样性 总被引:15,自引:1,他引:14
应用双梯度-变性梯度凝胶电泳(DG-DGGE)对生物制氢反应器微生物种群的动态变化及多样性进行监测。间隔7d从反应器取厌氧活性污泥,以细菌16SrDNA通用引物进行V2~V3区域PCR扩增,长约450bp的PCR产物经DGGE分离后,获得污泥微生物群落的16SrDNA指纹图谱。污泥接种到反应器后微生物群落中既有原始种群的消亡和增长,也有次级种群的强化和演变。反应器在运行初期群落演替迅速,15d时微生物群落结构变化最大。群落结构的相似性随着演替时间的增加而逐渐升高,种群动态变化后形成稳定的群落结构。29d时微生物多样性基本保持不变,微生物优势种属达到19个OTU。在细菌竞争和协同作用制约下,种群多样性降低后趋于稳定,形成顶级群落。有些种群在群落结构中一直存在,是群落建成的原始种群,原始种群与次级种群在代谢过程中具有协同作用,表现出群落的综合生态特征。 相似文献
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Observation of bias associated with re-amplification of DNA isolated from denaturing gradient gels 总被引:1,自引:0,他引:1
DNA from environmental PCR products separated by denaturing gradient gel electrophoresis (DGGE) was isolated from the background smear rather than from discrete bands of the DGGE gel. The "interband" region was considered as a potential source of less dominant members of natural microbial communities. Surprisingly, instead of detecting new bands from the re-amplified PCR products, patterns very similar to the original ones were obtained regardless of the position of the "interband" region. The results suggest that the separation of amplicons by DGGE may not be perfect and band re-amplification based sequence analyses need careful interpretation. 相似文献