Denaturing Gradient Gel Electrophoresis Analysis of Archaeal and Bacterial Populations in a Submerged Anaerobic Membrane Bioreactor Treating Landfill Leachate at Low Temperatures

2012 ◽  
Vol 29 (4) ◽  
pp. 219-226 ◽  
Author(s):  
Antoine P. Trzcinski ◽  
David C. Stuckey
Keyword(s):  
Landfill Leachate ◽  
Low Temperatures ◽  
Gel Electrophoresis ◽  
Membrane Bioreactor ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Populations ◽  
Anaerobic Membrane Bioreactor ◽  
Gradient Gel Electrophoresis

scholarly journals Design and Evaluation of PCR Primers for Analysis of Bacterial Populations in Wine by Denaturing Gradient Gel Electrophoresis

2003 ◽  
Vol 69 (11) ◽  
pp. 6801-6807 ◽  
Author(s):  
Isabel Lopez ◽  
Fernanda Ruiz-Larrea ◽  
Luca Cocolin ◽  
Erica Orr ◽  
Trevor Phister ◽  
...  
Keyword(s):  
Lactic Acid ◽  
16S Rdna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Species ◽  
Pcr Primers ◽  
Fungal Species ◽  
Bacterial Populations ◽  
Gradient Gel Electrophoresis ◽  
Primer Sets

ABSTRACT Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified ribosomal DNA (rDNA) is routinely used to compare levels of diversity of microbial communities and to monitor population dynamics. While using PCR-DGGE to examine the bacteria in wine fermentations, we noted that several commonly used PCR primers for amplifying bacterial 16S rDNA also coamplified yeast, fungal, or plant DNA present in samples. Unfortunately, amplification of nonbacterial DNA can result in a masking of bacterial populations in DGGE profiles. To surmount this problem, we developed two new primer sets for specific amplification of bacterial 16S rDNA in wine fermentation samples without amplification of eukaryotic DNA. One primer set, termed WLAB1 and WLAB2, amplified lactic acid bacteria, while another, termed WBAC1 and WBAC2, amplified both lactic acid bacterial and acetic acid bacterial populations found in wine. Primer specificity and efficacy were examined with DNA isolated from numerous bacterial, yeast, and fungal species commonly found in wine and must samples. Importantly, both primer sets effectively distinguished bacterial species in wine containing mixtures of yeast and bacteria.

Denaturing gradient gel electrophoresis analysis of a methyl tert-butyl ether degrading culture applied in a membrane bioreactor

2002 ◽  
Vol 2 (2) ◽  
pp. 207-212 ◽  
Author(s):  
A. Pruden ◽  
M. Suidan ◽  
J. Morrison ◽  
A. Venosa
Keyword(s):  
Gel Electrophoresis ◽  
Membrane Bioreactor ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Selective Pressure ◽  
Methyl Tert Butyl Ether ◽  
Butyl Ether ◽  
Tert Butyl ◽  
Start Up ◽  
Gradient Gel Electrophoresis ◽  
Dramatic Shift

A membrane bioreactor (MBR) was operated for the removal of methyl tert-butyl ether (MTBE) from water. Although the reactor was seeded with several cultures acclimated to MTBE degradation, a long start-up time was observed. Monitoring of the reactor with denaturing gradient gel electrophoresis (DGGE) revealed a dramatic shift in the MBR culture from the original seed culture, indicating that the membrane had exerted a selective pressure on the culture. The MBR culture was found to be dominated almost entirely by Sphingomonas, belonging to the a-4 subclass of the a-Proteobacteria. Several unique properties of Sphingomonas, including their characteristic outer membrane containing glycosphingolipids, as well as their extreme adeptness at xenobiotic degradation are hypothesized to have aided in their selection in this bioreactor.

scholarly journals Analysis of Bacterial Communities in the Rhizosphere of Chrysanthemum via Denaturing Gradient Gel Electrophoresis of PCR-Amplified 16S rRNA as Well as DNA Fragments Coding for 16S rRNA

2001 ◽  
Vol 67 (1) ◽  
pp. 172-178 ◽  
Author(s):  
Bernadette M. Duineveld ◽  
George A. Kowalchuk ◽  
Anneke Keijzer ◽  
Jan Dirk van Elsas ◽  
Johannes A. van Veen
Keyword(s):  
16S Rrna ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bulk Soil ◽  
Root Tip ◽  
Rt Pcr ◽  
Root Tips ◽  
Bacterial Populations ◽  
Gradient Gel Electrophoresis ◽  
Species Specific

ABSTRACT The effect of developing chrysanthemum roots on the presence and activity of bacterial populations in the rhizosphere was examined by using culture-independent methods. Nucleic acids were extracted from rhizosphere soil samples associated with the bases of roots or root tips of plants harvested at different stages of development. PCR and reverse transcriptase (RT) PCR were used to amplify 16S ribosomal DNA (rDNA) and 16S rRNA, respectively, and the products were subjected to denaturing gradient gel electrophoresis (DGGE). Prominent DGGE bands were excised and sequenced to gain insight into the identities of predominantly present (PCR) and predominantly active (RT-PCR) bacterial populations. The majority of DGGE band sequences were related to bacterial genera previously associated with the rhizosphere, such asPseudomonas, Comamonas, Variovorax, and Acetobacter, or typical of root-free soil environments, such as Bacillus and Arthrobacter. The PCR-DGGE patterns observed for bulk soil were somewhat more complex than those obtained from rhizosphere samples, and the latter contained a subset of the bands present in bulk soil. DGGE analysis of RT-PCR products detected a subset of bands visible in the rDNA-based analysis, indicating that some dominantly detected bacterial populations did not have high levels of metabolic activity. The sequences detected by the RT-PCR approach were, however, derived from a wide taxonomic range, suggesting that activity in the rhizosphere was not determined at broad taxonomic levels but rather was a strain- or species-specific phenomenon. Comparative analysis of DGGE profiles grouped all DNA-derived root tip samples together in a cluster, and within this cluster the root tip samples from young plants formed a separate subcluster. Comparison of rRNA-derived bacterial profiles showed no grouping of root tip samples versus root base samples. Rather, all profiles derived from 2-week-old plant rhizosphere soils grouped together regardless of location along the root.

scholarly journals Denaturing Gradient Gel Electrophoresis Analysis of 16S Ribosomal DNA Amplicons To Monitor Changes in Fecal Bacterial Populations of Weaning Pigs after Introduction of Lactobacillus reuteri Strain MM53

2000 ◽  
Vol 66 (11) ◽  
pp. 4705-4714 ◽  
Author(s):  
Joyce M. Simpson ◽  
Vance J. McCracken ◽  
H. Rex Gaskins ◽  
Roderick I. Mackie
Keyword(s):  
16S Rdna ◽  
Gel Electrophoresis ◽  
Lactobacillus Reuteri ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Populations ◽  
Variable Regions ◽  
Select Group ◽  
Genes Encoding ◽  
Gradient Gel Electrophoresis ◽  
Weaning Pigs

ABSTRACT The diversity and stability of the fecal bacterial microbiota in weaning pigs was studied after introduction of an exogenousLactobacillus reuteri strain, MM53, using a combination of cultivation and techniques based on genes encoding 16S rRNA (16S rDNA). Piglets (n = 9) were assigned to three treatment groups (control, daily dosed, and 4th-day dosed), and fresh fecal samples were collected daily. Dosed animals received 2.5 × 1010 CFU of antibiotic-resistant L. reuteriMM53 daily or every 4th day. Mean Lactobacillus counts for the three groups ranged from 1 × 109 to 4 × 109 CFU/g of feces. Enumeration of strain L. reuteri MM53 on MRS agar (Difco) plates containing streptomycin and rifampin showed that the introduced strain fluctuated between 8 × 103 and 5 × 106 CFU/g of feces in the two dosed groups. Denaturing gradient gel electrophoresis (DGGE) of PCR-amplified 16S rDNA fragments, with primers specific for variable regions 1 and 3 (V1 and V3), was used to profile complexity of fecal bacterial populations. Analysis of DGGE banding profiles indicated that each individual maintained a unique fecal bacterial population that was stable over time, suggesting a strong host influence. In addition, individual DGGE patterns could be separated into distinct time-dependent clusters. Primers designed specifically to restrict DGGE analysis to a select group of lactobacilli allowed examination of interspecies relationships and abundance. Based on relative band migration distance and sequence determination, L. reuteriwas distinguishable within the V1 region 16S rDNA gene patterns. Daily fluctuations in specific bands within these profiles were observed, which revealed an antagonistic relationship between L. reuteri MM53 (band V1-3) and another indigenousLactobacillus assemblage (band V1-6).

scholarly journals Monitoring of the succession of bacterial populations in infant faeces by PCR-denaturing gradient gel electrophoresis

2005 ◽  
Vol 17 (4) ◽  
pp. 205-211 ◽  
Author(s):  
M. José Pozuelo De Felipe ◽  
Raimundo García-Albiach ◽  
Alejandra Montesi Libois ◽  
Coronación Rodríguez Borrajo ◽  
Rosa Del Campo ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Populations ◽  
Gradient Gel Electrophoresis

Monitoring of the succession of bacterial populations in infant faeces by PCR-denaturing gradient gel electrophoresis

2005 ◽  
Vol 17 (4) ◽  
Author(s):  
M. José Pozuelo De Felipe ◽  
Raimundo García-Albiach ◽  
Alejandra Montesi Libois ◽  
Coronacion Rodríguez Borrajo ◽  
Rosa Del Campo ◽  
...  
Keyword(s):  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Bacterial Populations ◽  
Gradient Gel Electrophoresis

scholarly journals Isolation of Methylophaga spp. from Marine Dimethylsulfide-Degrading Enrichment Cultures and Identification of Polypeptides Induced during Growth on Dimethylsulfide

2007 ◽  
Vol 73 (8) ◽  
pp. 2580-2591 ◽  
Author(s):  
Hendrik Schäfer
Keyword(s):  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Large Subunit ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Butyl Ether ◽  
Bacterial Populations ◽  
Enrichment Cultures ◽  
Gradient Gel Electrophoresis ◽  
Insight Into

ABSTRACT Dimethylsulfide (DMS)-degrading enrichment cultures were established from samples of coastal seawater, nonaxenic Emiliania huxleyi cultures, and mixed marine methyl halide-degrading enrichment cultures. Bacterial populations from a broad phylogenetic range were identified in the mixed DMS-degrading enrichment cultures by denaturing gradient gel electrophoresis (DGGE). Sequences of dominant DGGE bands were similar to those of members of the genera Methylophaga and Alcanivorax. Several closely related Methylophaga strains were obtained that were able to grow on DMS as the carbon and energy source. Roseobacter-related populations were detected in some of the enrichment cultures; however, none of the Roseobacter group isolates that were tested were able to grow on DMS. Oxidation of DMS by Methylophaga sp. strain DMS010 was not affected by addition of the inhibitor chloroform or methyl tert-butyl ether, suggesting that DMS metabolism may occur by a route different from those described for Thiobacillus species and other unidentified marine isolates. Addition of DMS and methanethiol to whole-cell suspensions of strain DMS010 induced oxygen uptake when strain DMS010 was grown on DMS but not in cells grown on methanol. The apparent Km s of strain DMS010 for DMS and for methanethiol were 2.1 and 4.6 μM, respectively, when grown on DMS. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the biomass of strain DMS010 and analysis of peptide bands by mass spectrometry techniques and N-terminal sequencing provided the first insight into the identity of polypeptides induced during growth on DMS. These included XoxF, a homolog of the large subunit of methanol dehydrogenase for which a biological role has not been identified previously.

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