Online citations, reference lists, and bibliographies.

Bias Caused By Template Annealing In The Amplification Of Mixtures Of 16S RRNA Genes By PCR.

M. Suzuki, S. Giovannoni
Published 1996 · Medicine, Biology
Cite This
Download PDF
Analyze on Scholarcy
The PCR is used widely for the study of rRNA genes amplified from mixed microbial populations. These studies resemble quantitative applications of PCR in that the templates are mixtures of homologs and the relative abundance of amplicons is thought to provide some measure of the gene ratios in the starting mixture. Although such studies have established the presence of novel rRNA genes in many natural ecosystems, inferences about gene abundance have been limited by uncertainties about the relative efficiency of gene amplification in the PCR. To address this question, three rRNA gene standards were prepared by PCR, mixed in known proportions, and amplified a second time by using primer pairs in which one primer was labeled with a fluorescent nucleotide derivative. The PCR products were digested with restriction endonucleases, and the frequencies of genes in the products were determined by electrophoresis on an Applied Biosystems 373A automated DNA sequencer in Genescan mode. Mixtures of two templates amplified with the 519F-1406R primer pair yielded products in the predicted proportions. A second primer pair (27F-338R) resulted in strong bias towards 1:1 mixtures of genes in final products, regardless of the initial proportions of the templates. This bias was strongly dependent on the number of cycles of replication. The results fit a kinetic model in which the reannealing of genes progressively inhibits the formation of template-primer hybrids.
This paper references
The impact of the PCR plateau phase on quantitative PCR.
Ciaran G. Morrison (1994)
Molecular cloning: A laboratory manual 2nd ed. Edited by J. . Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989 (in 3 volumes)
P. Deininger (1990)
Survival and activity of Pseudomonas sp. strain B13(FR1) in a marine microcosm determined by quantitative PCR and an rRNA-targeting probe and its effect on the indigenous bacterioplankton.
Thomas Dyrmann Leser (1995)
PCR : the first few cycles
E. F. Fritsch (1991)
Analysis of cytokine mRNA and DNA: detection and quantitation by competitive polymerase chain reaction.
G. Gilliland (1990)
Plateau effect — understanding PCR limitations
P. D. Siebert (1993)
PCR: the first few cycles. Amplifications 7:1–4
G. Ruano (1991)
Competitive PCR
P. Siebert (1992)
[The polymerase chain reaction].
T. Arai (1991)
Phylogenetic diversity of subsurface marine microbial communities from the Atlantic and Pacific oceans.
J. Fuhrman (1993)
A novel procedure for quantitative polymerase chain reaction by coamplification of competitive templates.
S. Diviacco (1992)
Quantitation of mRNA by the polymerase chain reaction.
A. M. Wang (1989)
Differential amplification of rRNA genes by polymerase chain reaction.
A. L. Reysenbach (1992)
A commentary on the practical applications of competitive PCR.
L. Raeymaekers (1995)
Surveys of Gene Families Using Polymerase Chain Reaction: PCR Selection and PCR Drift
A. Wagner (1994)
Quantitative PCR: Procedures and precisions
Jerry Nedelman (1992)
Phylogenetic analysis of a natural marine bacterioplankton population by rRNA gene cloning and sequencing.
T. B. Britschgi (1991)
Phylogenetic diversity of aggregate‐attached vs. free‐living marine bacterial assemblages
E. DeLong (1993)
Quantitative PCR: theoretical considerations with practical implications.
L. Raeymaekers (1993)
General method for amplifying regions of very high G+C content.
C. Dutton (1993)
Absolute mRNA quantification using the polymerase chain reaction (PCR). A novel approach by a PCR aided transcript titration assay (PATTY).
M. Becker-André (1989)
Plateau effect-understanding PCR limitations
Alessandro Sardelli (1993)
Profiling complex microbial populations by denaturing gradient gel electrophoresis analysis of polym
Gerard Muyzer (1993)
Competitive PCR. Nature (London) 359:557–558
P. D. Siebert (1992)
A molecular technique for identification of bacteria using small subunit ribosomal RNA sequences.
E. Avaniss-Aghajani (1994)

This paper is referenced by
Soil Fungal Communities Underneath Willow Canopies on a Primary Successional Glacier Forefront: rDNA Sequence Results Can Be Affected by Primer Selection and Chimeric Data
A. Jumpponen (2004)
Volatilization and Precipitation of Tellurium by Aerobic, Tellurite-Resistant Marine Microbes
Patrick R. L. Ollivier (2008)
In Situ Biostimulation of Uranium Reducing Microorganisms at the Old Rifle UMTRA Site
Y. Chang (2005)
Probiotic bacteria for hatchery production of Greenshell mussels, Perna canaliculus
Aditya Kesarcodi-Watson (2009)
Archaeal communities associated with shallow to deep subseafloor sediments of the New Caledonia Basin.
Erwan G. Roussel (2009)
Real-time polymerase chain reaction quantification of Porphyromonas gingivalis and Tannerella forsythia in primary endodontic infections.
Daniel Saito (2009)
Two electrophoreses in different pH buffers to purify forest soil DNA contaminated with humic substances
Weiguo Hou (2010)
Molecular Source Tracking of Bioaerosols in the Quarantined Katrina Flood Zone
Mari Rodríguez de Evgrafov (2010)
Advancing the understanding of biogeography-diversity relationships of benthic microorganisms in the North Sea.
Melanie Sapp (2010)
Analysis of bacterial diversity in the intestine of grass carp (Ctenopharyngodon idellus) based on 16S rDNA gene sequences
Shaofeng Han (2010)
First report of diazotrophic Brevundimonas spp. as growth enhancer and root colonizer of potato
T. Naqqash (2020)
Microbiomes of ant castes implicate new microbial roles in the fungus-growing ant Trachymyrmex septentrionalis
Heather D. Ishak (2011)
Comparison of DNA extraction methodologies used for assessing fungal diversity via ITS sequencing.
William R. Rittenour (2012)
Do diet and taxonomy influence insect gut bacterial communities?
Daniel R. Colman (2012)
Comparative metagenomic and rRNA microbial diversity characterization using archaeal and bacterial synthetic communities.
Migun Shakya (2013)
Plant Growth Promotion Potential Is Equally Represented in Diverse Grapevine Root-Associated Bacterial Communities from Different Biopedoclimatic Environments
Ramona Marasco (2013)
Protocols for Investigating the Microbiology of Coal-Bed-Produced Waters
Amy V. Callaghan (2014)
Microbial community structure and metabolic property of biofilms in vermifiltration for liquid-state sludge stabilization using PLFA profiles.
Chunhui Zhao (2014)
Community Analysis of Ammonia Oxidizing Bacteria through MolecularGenetics in Activated Sludge of Effluent Treatment Plant
Maulin Pramod Shah (2015)
Diversity and abundance of "Pelagibacterales" (SAR11) in the Baltic Sea salinity gradient.
Daniel P. R. Herlemann (2014)
New lactic acid bacterial strains from traditional Mongolian fermented milk products have altered adhesion to porcine gastric mucin depending on the carbon source.
Hiromi Kimoto-Nira (2015)
Revisiting N2 fixation in Guerrero Negro intertidal microbial mats with a functional single-cell approach
Dagmar Woebken (2015)
Diversity of freshwater Epsilonproteobacteria and dark inorganic carbon fixation in the sulphidic redoxcline of a meromictic karstic lake.
Imma Noguerola (2015)
Phylogenetically Diverse Aerobic Anoxygenic Phototrophic Bacteria Isolated from Epilithic Biofilms in Tama River, Japan
Setsuko Hirose (2016)
A novel ultra high-throughput 16S rRNA amplicon sequencing library preparation method on the Illumina HiSeq platform
Eric Jacques de Muinck (2017)
A novel ultra high-throughput 16S rRNA gene amplicon sequencing library preparation method for the Illumina HiSeq platform
Eric Jacques de Muinck (2017)
Active Archaeal Communities at Cold Seep Sediments Populated by Siboglinidae Tubeworms from the Storegga Slide
Cassandre Sara Lazar (2010)
Phylogenetic Analysis of Bacteria Preserved in a Permafrost Ice Wedge for 25,000 Years
T. Katayama (2007)
Patterns and governing forces in aquatic microbial communities
S. Nold (2004)
Evidence of Purifying Selection and Co-Evolution at the Fold-Back Arm of the Novel Precursor MicroRNA159 Gene in Phalaenopsis Species (Orchidaceae)
Chi-Chu Tsai (2014)
Differential diagnosis of tick-borne diseases and population genetic analysis of Babesia bovis and Babesia bigemina.
Martin Simuunza (2009)
The Cyanobacterial Biota of Polar Regions: a Molecular Approach
Igor Stelmach Pessi (2017)
See more
Semantic Scholar Logo Some data provided by SemanticScholar