Upstream-independent ribosomal RNA amplification analysis (URA): a new approach to characterizing the diversity of natural microbial communities

2001 ◽  
Vol 3 (10) ◽  
pp. 662-666 ◽  
Author(s):  
Michail M. Yakimov ◽  
Laura Giuliano ◽  
Kenneth N. Timmis ◽  
Peter N. Golyshin
Keyword(s):  
Microbial Communities ◽  
Ribosomal Rna ◽  
Rna Amplification ◽  
New Approach

scholarly journals Microarray-Based Analysis of Microbial Community RNAs by Whole-Community RNA Amplification

2006 ◽  
Vol 73 (2) ◽  
pp. 563-571 ◽  
Author(s):  
Haichun Gao ◽  
Zamin K. Yang ◽  
Terry J. Gentry ◽  
Liyou Wu ◽  
Christopher W. Schadt ◽  
...  
Keyword(s):  
Microbial Communities ◽  
Environmental Samples ◽  
Bacterial Species ◽  
Shewanella Oneidensis ◽  
Rna Amplification ◽  
T7 Promoter ◽  
New Approach ◽  
Functional Activities ◽  
Amplification Method ◽  
Groundwater Samples

ABSTRACT A new approach, termed whole-community RNA amplification (WCRA), was developed to provide sufficient amounts of mRNAs from environmental samples for microarray analysis. This method employs fusion primers (six to nine random nucleotides with an attached T7 promoter) for the first-strand synthesis. The shortest primer (T7N6S) gave the best results in terms of the yield and representativeness of amplification. About 1,200- to 1,800-fold amplification was obtained with amounts of the RNA templates ranging from 10 to 100 ng, and very representative detection was obtained with 50 to 100 ng total RNA. Evaluation with a Shewanella oneidensis Δfur strain revealed that the amplification method which we developed could preserve the original abundance relationships of mRNAs. In addition, to determine whether representative detection of RNAs can be achieved with mixed community samples, amplification biases were evaluated with a mixture containing equal quantities of RNAs (100 ng each) from four bacterial species, and representative amplification was also obtained. Finally, the method which we developed was applied to the active microbial populations in a denitrifying fluidized bed reactor used for denitrification of contaminated groundwater and ethanol-stimulated groundwater samples for uranium reduction. The genes expressed were consistent with the expected functions of the bioreactor and groundwater system, suggesting that this approach is useful for analyzing the functional activities of microbial communities. This is one of the first demonstrations that microarray-based technology can be used to successfully detect the activities of microbial communities from real environmental samples in a high-throughput fashion.

scholarly journals Molecular Approaches to Studying Microbial Communities: Targeting the 16S Ribosomal RNA Gene

2016 ◽  
Vol 38 (3) ◽  
pp. 223-232 ◽  
Author(s):  
Kazumasa FUKUDA ◽  
Midori OGAWA ◽  
Hatsumi TANIGUCHI ◽  
Mitsumasa SAITO
Keyword(s):  
Microbial Communities ◽  
Ribosomal Rna ◽  
16S Ribosomal Rna ◽  
Approaches To Studying ◽  
Ribosomal Rna Gene ◽  
Molecular Approaches ◽  
16S Ribosomal Rna Gene

scholarly journals MAPseq: improved speed, accuracy and consistency in ribosomal RNA sequence analysis

2017 ◽  
Author(s):  
João F Matias Rodrigues ◽  
Thomas SB Schmidt ◽  
Janko Tackmann ◽  
Christian von Mering
Keyword(s):  
Sequence Analysis ◽  
Microbial Communities ◽  
Study Design ◽  
Ribosomal Rna ◽  
Source Code ◽  
Shotgun Sequencing ◽  
Metagenomic Sequencing ◽  
Rna Sequence ◽  
Taxonomic Analysis ◽  
Speed Accuracy

AbstractMetagenomic sequencing has become crucial to studying microbial communities, but meaningful taxonomic analysis and inter-comparison of such data are still hampered by technical limitations, between-study design variability and inconsistencies between taxonomies used. Here we present MAPseq, a framework for reference-based rRNA metagenomic analysis that is up to 30% more accurate (F1/2 score) and up to one hundred times faster than existing solutions, providing in a single run multiple taxonomy classifications and hierarchical OTU mappings, for both amplicon and shotgun sequencing strategies, and for datasets of virtually any size. Availability: Source code and binaries are freely available at http://meringlab.org/software/mapseq/

Use of ribosomal RNA-based molecular probes for characterization of complex microbial communities in anaerobic biofilms

1995 ◽  
Vol 31 (1) ◽  
pp. 261-272 ◽  
Author(s):  
Lutgarde Raskin ◽  
Rudolf I. Amann ◽  
Lars K. Poulsen ◽  
Bruce E. Rittmann ◽  
David A. Stahl
Keyword(s):  
Microbial Communities ◽  
Ribosomal Rna ◽  
Fixed Bed ◽  
Molecular Probes ◽  
Biofilm Reactor ◽  
Sulfate Reducing ◽  
Biofilm Reactors ◽  
Functional Behavior ◽  
Long Term Study

The use of ribosomal RNA (rRNA) probe technology for the characterization of complex microbial communities is reviewed and illustrated by discussing the results of a long-term study of four anaerobic fixed-bed biofilm reactors. Two distinct approaches were used to characterize the microbial community structure in these biofilm reactors. The first used a collection of phylogenetically defined oligonucleotide rRNA probes for methanogens and sulfate-reducing bacteria (SRB) to quantify their populations. Population abundance was linked to the functional behavior of the biofilm reactor community by determining the effluent concentrations of the substrates, intermediates, and final products of microbial metabolism. This analysis indicated that the presence of SRB (especially Desulfovibrio-species) was not dependent upon the presence of sulfate. Methanobacteriales-species were the major competitors for hydrogen with these SRB in the absence of sulfate. The second approach involved selective amplification, cloning, sequencing, and whole cell hybridization to identify, visualize, and isolate a biofilm community member (strain PT-2). Subsequently, it was determined that the growth rate of strain PT-2 was significantly higher in young biofilms than in established biofilms.

An oligonucleotide analog approach to the decoding region of 16S rRNA

1995 ◽  
Vol 73 (11-12) ◽  
pp. 899-905 ◽  
Author(s):  
Seth Stern ◽  
Prakash Purohit
Keyword(s):  
16S Rrna ◽  
Ribosomal Rna ◽  
Molecular Basis ◽  
Ribosomal Subunit ◽  
Small Subunit ◽  
16S Ribosomal Rna ◽  
New Approach ◽  
16S Rna ◽  
Analog Approach ◽  
Decoding Region

Despite the passage of about 30 years since the discovery of the translational activities of ribosomes and the outlining of the roles of the large and small subunits, the actual molecular basis for the mRNA decoding activities of the small subunit has remained essentially obscure. In this paper, we describe a new approach using oligonucleotide analogs of 16S ribosomal RNA, in which the small ribosomal subunit is effectively deconstructed into a smaller more experimentally tractable form. Specifically, we review the results of experiments using an oligonucleotide analog of the decoding region of 16S ribosomal RNA, suggesting that the decoding region is the functional core of the small subunit, that it contacts both mRNA codons and tRNA anticodons, and that it mediates and probably enhances codon–anticodon base pairing, that is, decoding.Key words: translation, ribosome, 30S, 16S, RNA, decoding, antibiotic.

scholarly journals Microbial communities of poultry house dust, excreta and litter are partially representative of microbiota of chicken caecum and ileum

PLoS ONE ◽  
2021 ◽  
Vol 16 (8) ◽  
pp. e0255633
Author(s):  
Yugal R. Bindari ◽  
Robert J. Moore ◽  
Thi Thu Hao Van ◽  
Matthew Hilliar ◽  
Shu-Biao Wu ◽  
...  
Keyword(s):  
Microbial Communities ◽  
Ribosomal Rna ◽  
Sampling Methods ◽  
Amplicon Sequencing ◽  
Microbial Composition ◽  
Poultry House ◽  
Operational Taxonomic Units ◽  
Non Invasive ◽  
16S Ribosomal Rna Gene ◽  
Poultry Dust

Traditional sampling methods for the study of poultry gut microbiota preclude longitudinal studies as they require euthanasia of birds for the collection of caecal and ileal contents. Some recent research has investigated alternative sampling methods to overcome this issue. The main goal of this study was to assess to what extent the microbial composition of non-invasive samples (excreta, litter and poultry dust) are representative of invasive samples (caecal and ileal contents). The microbiota of excreta, dust, litter, caecal and ileal contents (n = 110) was assessed using 16S ribosomal RNA gene amplicon sequencing. Of the operational taxonomic units (OTUs) detected in caecal contents, 99.7% were also detected in dust, 98.6% in litter and 100% in excreta. Of the OTUs detected in ileal contents, 99.8% were detected in dust, 99.3% in litter and 95.3% in excreta. Although the majority of the OTUs found in invasive samples were detected in non-invasive samples, the relative abundance of members of the microbial communities of these groups were different, as shown by beta diversity measures. Under the conditions of this study, correlation analysis showed that dust could be used as a proxy for ileal and caecal contents to detect the abundance of the phylum Firmicutes, and excreta as a proxy of caecal contents for the detection of Tenericutes. Similarly, litter could be used as a proxy for caecal contents to detect the abundance of Firmicutes and Tenericutes. However, none of the non-invasive samples could be used to infer the overall abundance of OTUs observed in invasive samples. In conclusion, non-invasive samples could be used to detect the presence and absence of the majority of the OTUs found in invasive samples, but could not accurately reflect the microbial community structure of invasive samples.

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