scholarly journals Improved group-specific PCR primers for denaturing gradient gel electrophoresis analysis of the genetic diversity of complex microbial communities

2008 ◽  
Vol 2 (4) ◽  
pp. 379-392 ◽  
Author(s):  
Martin Mühling ◽  
John Woolven-Allen ◽  
J Colin Murrell ◽  
Ian Joint
Keyword(s):  
Genetic Diversity ◽  
Microbial Communities ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Pcr Primers ◽  
Specific Pcr ◽  
Gradient Gel Electrophoresis

scholarly journals Microbial Community Fingerprinting by Differential Display-Denaturing Gradient Gel Electrophoresis

2010 ◽  
Vol 77 (1) ◽  
pp. 351-354 ◽  
Author(s):  
M. C. Portillo ◽  
D. Villahermosa ◽  
A. Corzo ◽  
J. M. Gonzalez
Keyword(s):  
Microbial Community ◽  
Microbial Communities ◽  
Differential Display ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Pcr Primers ◽  
Gradient Gel Electrophoresis ◽  
Community Fingerprinting ◽  
Community Complexity ◽  
Partial Fingerprint

ABSTRACTComplex microbial communities exhibit a large diversity, hampering differentiation by DNA fingerprinting. Herein, differential display-denaturing gradient gel electrophoresis is proposed. By adding a nucleotide to the 3′ ends of PCR primers, 16 primer pairs and fingerprints were generated per community. Complexity reduction in each partial fingerprint facilitates sample comparison.

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 The effect of inoculum dose on the genetic diversity detected within Helicoverpa armigera nucleopolyhedrovirus populations

2013 ◽  
Vol 94 (11) ◽  
pp. 2524-2529 ◽  
Author(s):  
Vicky Lynne Baillie ◽  
Gustav Bouwer
Keyword(s):  
Genetic Diversity ◽  
Helicoverpa Armigera ◽  
Gel Electrophoresis ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Model System ◽  
Inoculum Dose ◽  
Significant Difference ◽  
Helicoverpa Armígera ◽  
Gradient Gel Electrophoresis ◽  
Neonate Larvae

Environmental and infection variables may affect the genetic diversity of baculovirus populations. In this study, Helicoverpa armigera nucleopolyhedrovirus (HearNPV) was used as a model system for studying the effects of a key infection variable, inoculum dose, on the genetic diversity within nucleopolyhedrovirus populations. Diversity and equitability indices were calculated from DNA polymerase-specific denaturing gradient gel electrophoresis profiles obtained from individual H. armigera neonate larvae inoculated with either an LD5 or LD95 of HearNPV. Although the genetic diversity detected in larvae treated with an LD95 was not statistically different from the diversity detected in the HearNPV inoculum samples, there was a statistically significant difference in the genetic diversity detected in the LD5-inoculated larvae compared with the genetic diversity detected in the HearNPV samples used for the inoculations. The study suggests that inoculum dose needs to be considered carefully in experiments that evaluate HearNPV genetic diversity or in studies where differences in genetic diversity may have phenotypic consequences.

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