Comparison of microbial communities between normal and swollen canned soy sauces using nested PCR-denaturing gradient gel electrophoresis, HPLC and plate techniques

2014 ◽  
Vol 49 (11) ◽  
pp. 2499-2505 ◽  
Author(s):  
Lili Cheng ◽  
Weifeng Lin ◽  
Pan Li ◽  
Jianfei Huang ◽  
Lixin Luo
Keyword(s):  
Microbial Communities ◽  
Nested Pcr ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Gradient Gel Electrophoresis

scholarly journals Nested PCR-Denaturing Gradient Gel Electrophoresis Approach To Determine the Diversity of Sulfate-Reducing Bacteria in Complex Microbial Communities

2005 ◽  
Vol 71 (5) ◽  
pp. 2325-2330 ◽  
Author(s):  
Shabir A. Dar ◽  
J. Gijs Kuenen ◽  
Gerard Muyzer
Keyword(s):  
Microbial Communities ◽  
Nested Pcr ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Sulfate Reducing Bacteria ◽  
Comparative Sequence Analysis ◽  
Rrna Gene ◽  
Sulfate Reducing ◽  
Gradient Gel Electrophoresis ◽  
Reducing Bacteria

ABSTRACT 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.

scholarly journals Development and Validation of a Nested-PCR-Denaturing Gradient Gel Electrophoresis Method for Taxonomic Characterization of Bifidobacterial Communities

2003 ◽  
Vol 69 (11) ◽  
pp. 6380-6385 ◽  
Author(s):  
R. Temmerman ◽  
L. Masco ◽  
T. Vanhoutte ◽  
G. Huys ◽  
J. Swings
Keyword(s):  
Nested Pcr ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Temporal Changes ◽  
Rrna Gene ◽  
Specific Pcr ◽  
Gradient Gel Electrophoresis ◽  
Species Specific ◽  
Taxonomic Characterization

ABSTRACT The taxonomic characterization of a bacterial community is difficult to combine with the monitoring of its temporal changes. None of the currently available identification techniques are able to visualize a “complete” community, whereas techniques designed for analyzing bacterial ecosystems generally display limited or labor-intensive identification potential. This paper describes the optimization and validation of a nested-PCR-denaturing gradient gel electrophoresis (DGGE) approach for the species-specific analysis of bifidobacterial communities from any ecosystem. The method comprises a Bifidobacterium-specific PCR step, followed by purification of the amplicons that serve as template DNA in a second PCR step that amplifies the V3 and V6-V8 regions of the 16S rRNA gene. A mix of both amplicons is analyzed on a DGGE gel, after which the band positions are compared with a previously constructed database of reference strains. The method was validated through the analysis of four artificial mixtures, mimicking the possible bifidobacterial microbiota of the human and chicken intestine, a rumen, and the environment, and of two fecal samples. Except for the species Bifidobacterium coryneforme and B. indicum, all currently known bifidobacteria originating from various ecosystems can be identified in a highly reproducible manner. Because no further cloning and sequencing of the DGGE bands is necessary, this nested-PCR-DGGE technique can be completed within a 24-h span, allowing the species-specific monitoring of temporal changes in the bifidobacterial community.

scholarly journals Denaturing gradient gel electrophoresis as a fingerprinting method for the analysis of soil microbial communities

2009 ◽  
Vol 55 (No. 10) ◽  
pp. 413-423 ◽  
Author(s):  
V. Valášková ◽  
P. Baldrian
Keyword(s):  
Microbial Communities ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Soil Microbial Communities ◽  
Soil Microbial ◽  
Genes Encoding ◽  
Gradient Gel Electrophoresis ◽  
Temperature Gradient Gel Electrophoresis ◽  
Experimental Treatments ◽  
Multiple Samples

In soil microbial ecology, the effects of environmental factors and their gradients, temporal changes or the response to specific experimental treatments of microbial communities can only be effectively analyzed using methods that address the structural differences among whole communities. Fingerprinting methods are the most appropriate technique for this task when multiple samples must be analyzed. Among the methods currently used to compare microbial communities based on nucleic acid sequences, the techniques based on differences in the melting properties of double-stranded molecules, denaturing gradient gel electrophoresis (DGGE) or temperature gradient gel electrophoresis (TGGE), are the most widely used. Their main advantage is that they provide the possibility to further analyze whole sequences contained in fingerprints using molecular methods. In addition to the analysis of microbial communities based on DNA extracted from soils, DGGE/TGGE can also be used for the assessment of the active part of the community based on the analysis of RNA-derived sequences or for the analysis of sequences of functional genes encoding for proteins involved in important soil processes.

Spatial variation in microbial community structure, richness, and diversity in an alluvial aquifer

2012 ◽  
Vol 58 (9) ◽  
pp. 1135-1151 ◽  
Author(s):  
P.G. Medihala ◽  
J.R. Lawrence ◽  
G.D.W. Swerhone ◽  
D.R. Korber
Keyword(s):  
Community Structure ◽  
Microbial Community ◽  
Spatial Variability ◽  
Microbial Communities ◽  
Microbial Community Structure ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Microbial Community Composition ◽  
Alluvial Aquifer ◽  
Gradient Gel Electrophoresis

Relatively little is known regarding the spatial variability of microbial communities in aquifers where well fouling is an issue. In this study 2 water wells were installed in an alluvial aquifer located adjacent to the North Saskatchewan River and an associated piezometer network developed to facilitate the study of microbial community structure, richness, and diversity. Carbon utilization data analysis revealed reduced microbial activity in waters collected close to the wells. Functional PCR and quantitative PCR analysis indicated spatial variability in the potential for iron-, sulphate-, and nitrate-reducing activity at all locations in the aquifer. Denaturing gradient gel electrophoresis analysis of aquifer water samples using principal components analyses indicated that the microbial community composition was spatially variable, and denaturing gradient gel electrophoresis sequence analysis revealed that bacteria belonging to the genera Acidovorax , Rhodobacter , and Sulfuricurvum were common throughout the aquifer. Shannon’s richness (H′) and Pielou’s evenness (J′) indices revealed a varied microbial diversity (H′ = 1.488–2.274) and an even distribution of microbial communities within the aquifer (J′ = 0.811–0.917). Overall, these analyses revealed that the aquifer’s microbial community varied spatially in terms of composition, richness, and metabolic activity. Such information may facilitate the diagnosis, prevention, and management of fouling.

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