scholarly journals Rates of gene conversions between Escherichia coli ribosomal operons

2020 ◽  
Vol 11 (2) ◽  
Author(s):  
Isaac Gifford ◽  
Aurko Dasgupta ◽  
Jeffrey E Barrick
Keyword(s):  
Escherichia Coli ◽  
Gene Conversion ◽  
Accurate Estimate ◽  
Rrna Genes ◽  
Homologous Sequence ◽  
Rrna Gene ◽  
Multiple Copies ◽  
Ribosomal Operons ◽  
High Sequence Conservation ◽  
Gene Conversions

Abstract Due to their universal presence and high sequence conservation, ribosomal RNA (rRNA) sequences are used widely in phylogenetics for inferring evolutionary relationships between microbes and in metagenomics for analyzing the composition of microbial communities. Most microbial genomes encode multiple copies of rRNA genes to supply cells with sufficient capacity for protein synthesis. These copies typically undergo concerted evolution that keeps their sequences identical, or nearly so, due to gene conversion, a type of intragenomic recombination that changes one copy of a homologous sequence to exactly match another. Widely varying rates of rRNA gene conversion have previously been estimated by comparative genomics methods and using genetic reporter assays. To more directly measure rates of rRNA intragenomic recombination, we sequenced the seven Escherichia coli rRNA operons in 15 lineages that were evolved for ∼13,750 generations with frequent single-cell bottlenecks that reduce the effects of selection. We identified 38 gene conversion events and estimated an overall rate of intragenomic recombination within the 16S and 23S genes between rRNA copies of 3.6 × 10−4 per genome per generation or 8.6 × 10−6 per rRNA operon per homologous donor operon per generation. This rate varied only slightly from random expectations at different sites within the rRNA genes and between rRNA operons located at different positions in the genome. Our accurate estimate of the rate of rRNA gene conversions fills a gap in our quantitative understanding of how ribosomal sequences and other multicopy elements diversify and homogenize during microbial genome evolution.

scholarly journals Rates of gene conversions between Escherichia coli ribosomal operons

2020 ◽  
Author(s):  
Isaac Gifford ◽  
Aurko Dasgupta ◽  
Jeffrey E. Barrick
Keyword(s):  
Escherichia Coli ◽  
Gene Conversion ◽  
Accurate Estimate ◽  
Rrna Genes ◽  
Homologous Sequence ◽  
Rrna Gene ◽  
Potential Donor ◽  
Ribosomal Operons ◽  
High Sequence Conservation ◽  
Gene Conversions

ABSTRACTDue to their universal presence and high sequence conservation, rRNA sequences are used widely in phylogenetics for inferring evolutionary relationships between microbes and in metagenomics for analyzing the composition of microbial communities. Most microbial genomes encode multiple copies of ribosomal RNA (rRNA) genes to supply cells with sufficient capacity for protein synthesis. These copies typically undergo concerted evolution that keeps their sequences identical, or nearly so, due to gene conversion, a type of intragenomic recombination that changes one copy of a homologous sequence to exactly match another. Widely varying rates of rRNA gene conversion have previously been estimated by comparative genomics methods and using genetic reporter assays. To more directly measure rates of rRNA intragenomic recombination, we sequenced the seven Escherichia coli rRNA operons in 15 lineages of cells that were evolved for ~13,750 generations with frequent single-cell bottlenecks that reduce the effects of selection. We identified 34 gene conversion events and estimate an overall rate of intragenomic recombination events between rRNA copies of 3.2 × 10−4 per generation or 5.3 × 10−5 per potential donor sequence. This rate varied only slightly from random expectations between different portions of the rRNA genes and between rRNA operons located at different locations in the genome. This accurate estimate of the rate of rRNA gene conversions fills a gap in our quantitative understanding of how ribosomal sequences and other multicopy elements diversify and homogenize during microbial genome evolution.

scholarly journals Selective and Sensitive Method for PCR Amplification of Escherichia coli 16S rRNA Genes in Soil

2000 ◽  
Vol 66 (2) ◽  
pp. 844-849 ◽  
Author(s):  
G. Sabat ◽  
P. Rose ◽  
W. J. Hickey ◽  
J. M. Harkin
Keyword(s):  
Escherichia Coli ◽  
16S Rrna ◽  
Pcr Amplification ◽  
Pcr Primers ◽  
Enteric Bacteria ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Selective Amplification ◽  
E Coli

ABSTRACT A set of PCR primers targeting 16S rRNA gene sequences was designed, and PCR parameters were optimized to develop a robust and reliable protocol for selective amplification of Escherichia coli 16S rRNA genes. The method was capable of discriminatingE. coli from other enteric bacteria, including its closest relative, Shigella. Selective amplification of E. coli occurred only when the annealing temperature in the PCR was elevated to 72°C, which is 10°C higher than the optimum for the primers. Sensitivity was retained by modifying the length of steps in the PCR, by increasing the number of cycles, and most importantly by optimizing the MgCl2 concentration. The PCR protocol developed can be completed in less then 2 h and, by using Southern hybridization, has a detection limit of ca. 10 genomic equivalents per reaction. The method was demonstrated to be effective for detectingE. coli DNA in heterogeneous DNA samples, such as those extracted from soil.

scholarly journals Genome-Derived Criteria for Assigning Environmental narG and nosZ Sequences to Operational Taxonomic Units of Nitrate Reducers

2009 ◽  
Vol 75 (15) ◽  
pp. 5170-5174 ◽  
Author(s):  
Katharina Palmer ◽  
Harold L. Drake ◽  
Marcus A. Horn
Keyword(s):  
16S Rrna ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Operational Taxonomic Units ◽  
Nitrate Reducers ◽  
Multiple Copies ◽  
Tree Topologies ◽  
Gene Similarity ◽  
Very High

ABSTRACT Ninety percent of cultured bacterial nitrate reducers with a 16S rRNA gene similarity of ≥97% had a narG or nosZ similarity of ≥67% or ≥80%, respectively, suggesting that 67% and 80% could be used as standardized, conservative threshold similarity values for narG and nosZ, respectively (i.e., any two sequences that are less similar than the threshold similarity value have a very high probability of belonging to different species), for estimating species-level operational taxonomic units. Genus-level tree topologies of narG and nosZ were generally similar to those of the corresponding 16S rRNA genes. Although some genomes contained multiple copies of narG, recent horizontal gene transfer of narG was not apparent.

scholarly journals Intraspecific Diversity of the 23S rRNA Gene and the Spacer Region Downstream in Escherichia coli

1999 ◽  
Vol 181 (9) ◽  
pp. 2703-2709 ◽  
Author(s):  
Ana I. Antón ◽  
Antonio J. Martínez-Murcia ◽  
Francisco Rodríguez-Valera
Keyword(s):  
Escherichia Coli ◽  
Gene Families ◽  
Intraspecific Diversity ◽  
Rrna Genes ◽  
23S Rrna ◽  
Pcr Analysis ◽  
Rrna Gene ◽  
Enzyme Electrophoresis ◽  
Single Mutation ◽  
23S Rrna Gene

ABSTRACT The molecular microevolution of the 23S rRNA gene (rrl) plus the spacer downstream has been studied by sequencing of different operons from some representative strains of the Escherichia coli ECOR collection. The rrl gene was fully sequenced in six strains showing a total of 67 polymorphic sites, a level of variation per nucleotide similar to that found for the 16S rRNA gene (rrs) in a previous study. The size of the gene was highly conserved (2902 to 2905 nucleotides). Most polymorphic sites were clustered in five secondary-structure helices. Those regions in a large number of operons were sequenced, and several variations were found. Sequences of the same helix from two different strains were often widely divergent, and no intermediate forms existed. Intercistronic variability was detected, although it seemed to be lower than for the rrs gene. The presence of two characteristic sequences was determined by PCR analysis throughout all of the strains of the ECOR collection, and some correlations with the multilocus enzyme electrophoresis clusters were detected. The mode of variation of the rrl gene seems to be quite similar to that of therrs gene. Homogenization of the gene families and transfer of sequences from different clonal lines could explain this pattern of variation detected; perhaps these factors are more relevant to evolution than single mutation. The spacer region between the 23S and 5S rRNA genes exhibited a highly polymorphic region, particularly at the 3′ end.

Differentiation Of Clarias Batrachus, C. Gariepinus And Heteropneustes Fossilis By Pcr-Sequencing Of Mitochondrial 16s Rrna Gene

2015 ◽  
Vol 41 (1) ◽  
pp. 51-58
Author(s):  
Mohammad Shamimul Alam ◽  
Hawa Jahan ◽  
Rowshan Ara Begum ◽  
Reza M Shahjahan
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Fish Larva ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Valuable Insight ◽  
Multiple Sequence ◽  
Pcr Rflp ◽  
Alternative Means

Heteropneustesfossilis, Clariasbatrachus and C. gariepinus are three major catfishes ofecological and economic importance. Identification of these fish species becomes aproblem when the usual external morphological features of the fish are lost or removed,such as in canned fish. Also, newly hatched fish larva is often difficult to identify. PCRsequencingprovides accurate alternative means of identification of individuals at specieslevel. So, 16S rRNA genes of three locally collected catfishes were sequenced after PCRamplification and compared with the same gene sequences available from othergeographical regions. Multiple sequence alignment of the 16S rRNA gene fragments ofthe catfish species has revealed polymorphic sites which can be used to differentiate thesethree species from one another and will provide valuable insight in choosing appropriaterestriction enzymes for PCR-RFLP based identification in future. Asiat. Soc. Bangladesh, Sci. 41(1): 51-58, June 2015

scholarly journals Ecofriendly Synthesis of Silver Nanoparticles by Terrabacter humi sp. nov. and Their Antibacterial Application against Antibiotic-Resistant Pathogens

2020 ◽  
Vol 21 (24) ◽  
pp. 9746
Author(s):  
Shahina Akter ◽  
Sun-Young Lee ◽  
Muhammad Zubair Siddiqi ◽  
Sri Renukadevi Balusamy ◽  
Md. Ashrafudoulla ◽  
...  
Keyword(s):  
Silver Nanoparticles ◽  
Spherical Shape ◽  
Optimal Growth ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Strictly Aerobic ◽  
Drug Resistant ◽  
Protein Coding ◽  
A Genome ◽  
The Fourier Transform

It is essential to develop and discover alternative eco-friendly antibacterial agents due to the emergence of multi-drug-resistant microorganisms. In this study, we isolated and characterized a novel bacterium named Terrabacter humi MAHUQ-38T, utilized for the eco-friendly synthesis of silver nanoparticles (AgNPs) and the synthesized AgNPs were used to control multi-drug-resistant microorganisms. The novel strain was Gram stain positive, strictly aerobic, milky white colored, rod shaped and non-motile. The optimal growth temperature, pH and NaCl concentration were 30 °C, 6.5 and 0%, respectively. Based on 16S rRNA gene sequence, strain MAHUQ-38T belongs to the genus Terrabacter and is most closely related to several Terrabacter type strains (98.2%–98.8%). Terrabacter humi MAHUQ-38T had a genome of 5,156,829 bp long (19 contigs) with 4555 protein-coding genes, 48 tRNA and 5 rRNA genes. The culture supernatant of strain MAHUQ-38T was used for the eco-friendly and facile synthesis of AgNPs. The transmission electron microscopy (TEM) image showed the spherical shape of AgNPs with a size of 6 to 24 nm, and the Fourier transform infrared (FTIR) analysis revealed the functional groups responsible for the synthesis of AgNPs. The synthesized AgNPs exhibited strong anti-bacterial activity against multi-drug-resistant pathogens, Escherichia coli and Pseudomonas aeruginosa. Minimal inhibitory/bactericidal concentrations against E. coli and P. aeruginosa were 6.25/50 and 12.5/50 μg/mL, respectively. The AgNPs altered the cell morphology and damaged the cell membrane of pathogens. This study encourages the use of Terrabacter humi for the ecofriendly synthesis of AgNPs to control multi-drug-resistant microorganisms.

scholarly journals The Pif1 helicase is actively inhibited during meiotic recombination which restrains gene conversion tract length

2021 ◽  
Author(s):  
Dipti Vinayak Vernekar ◽  
Giordano Reginato ◽  
Céline Adam ◽  
Lepakshi Ranjha ◽  
Florent Dingli ◽  
...  
Keyword(s):  
Gene Conversion ◽  
Meiotic Recombination ◽  
Strand Break ◽  
Dna Helicase ◽  
Clamp Loader ◽  
Repair Synthesis ◽  
Dna Repair Synthesis ◽  
Gene Conversions

Abstract Meiotic recombination ensures proper chromosome segregation to form viable gametes and results in gene conversions events between homologs. Conversion tracts are shorter in meiosis than in mitotically dividing cells. This results at least in part from the binding of a complex, containing the Mer3 helicase and the MutLβ heterodimer, to meiotic recombination intermediates. The molecular actors inhibited by this complex are elusive. The Pif1 DNA helicase is known to stimulate DNA polymerase delta (Pol δ) -mediated DNA synthesis from D-loops, allowing long synthesis required for break-induced replication. We show that Pif1 is also recruited genome wide to meiotic DNA double-strand break (DSB) sites. We further show that Pif1, through its interaction with PCNA, is required for the long gene conversions observed in the absence of MutLβ recruitment to recombination sites. In vivo, Mer3 interacts with the PCNA clamp loader RFC, and in vitro, Mer3-MutLβ ensemble inhibits Pif1-stimulated D-loop extension by Pol δ and RFC-PCNA. Mechanistically, our results suggest that Mer3-MutLβ may compete with Pif1 for binding to RFC-PCNA. Taken together, our data show that Pif1’s activity that promotes meiotic DNA repair synthesis is restrained by the Mer3-MutLβ ensemble which in turn prevents long gene conversion tracts and possibly associated mutagenesis.

scholarly journals Microdiversity and phylogeographic diversification of bacterioplankton in pelagic freshwater systems revealed through long-read amplicon sequencing

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Yusuke Okazaki ◽  
Shohei Fujinaga ◽  
Michaela M. Salcher ◽  
Cristiana Callieri ◽  
Atsushi Tanaka ◽  
...  
Keyword(s):  
16S Rrna ◽  
Regional Scale ◽  
Scale Up ◽  
Amplicon Sequencing ◽  
Freshwater Ecosystems ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Long Read

Abstract Background Freshwater ecosystems are inhabited by members of cosmopolitan bacterioplankton lineages despite the disconnected nature of these habitats. The lineages are delineated based on > 97% 16S rRNA gene sequence similarity, but their intra-lineage microdiversity and phylogeography, which are key to understanding the eco-evolutional processes behind their ubiquity, remain unresolved. Here, we applied long-read amplicon sequencing targeting nearly full-length 16S rRNA genes and the adjacent ribosomal internal transcribed spacer sequences to reveal the intra-lineage diversities of pelagic bacterioplankton assemblages in 11 deep freshwater lakes in Japan and Europe. Results Our single nucleotide-resolved analysis, which was validated using shotgun metagenomic sequencing, uncovered 7–101 amplicon sequence variants for each of the 11 predominant bacterial lineages and demonstrated sympatric, allopatric, and temporal microdiversities that could not be resolved through conventional approaches. Clusters of samples with similar intra-lineage population compositions were identified, which consistently supported genetic isolation between Japan and Europe. At a regional scale (up to hundreds of kilometers), dispersal between lakes was unlikely to be a limiting factor, and environmental factors or genetic drift were potential determinants of population composition. The extent of microdiversification varied among lineages, suggesting that highly diversified lineages (e.g., Iluma-A2 and acI-A1) achieve their ubiquity by containing a consortium of genotypes specific to each habitat, while less diversified lineages (e.g., CL500-11) may be ubiquitous due to a small number of widespread genotypes. The lowest extent of intra-lineage diversification was observed among the dominant hypolimnion-specific lineage (CL500-11), suggesting that their dispersal among lakes is not limited despite the hypolimnion being a more isolated habitat than the epilimnion. Conclusions Our novel approach complemented the limited resolution of short-read amplicon sequencing and limited sensitivity of the metagenome assembly-based approach, and highlighted the complex ecological processes underlying the ubiquity of freshwater bacterioplankton lineages. To fully exploit the performance of the method, its relatively low read throughput is the major bottleneck to be overcome in the future.

scholarly journals 16S rRNA Gene Sequence Analysis ofPhotobacterium damselae and Nested PCR Method for Rapid Detection of the Causative Agent of Fish Pasteurellosis

1999 ◽  
Vol 65 (7) ◽  
pp. 2942-2946 ◽  
Author(s):  
Carlos R. Osorio ◽  
Matthew D. Collins ◽  
Alicia E. Toranzo ◽  
Juan L. Barja ◽  
Jesús L. Romalde
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Causative Agent ◽  
Nested Pcr ◽  
Gene Sequence ◽  
16S Rrna Gene Sequence ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Rrna Gene Sequence ◽  
Photobacterium Damselae

ABSTRACT The causative agent of fish pasteurellosis, the organism formerly known as Pasteurella piscicida, has been reclassified asPhotobacterium damselae subsp. piscicida on the basis of 16S rRNA gene sequence comparisons and chromosomal DNA-DNA hybridization data; thus, this organism belongs to the same species asPhotobacterium damselae subsp. damselae(formerly Vibrio damselae). Since reassignment of P. damselae subsp. piscicida was based on only two strains, one objective of the present work was to confirm the taxonomic position of this fish pathogen by sequencing the 16S rRNA genes of 26 strains having different geographic and host origins. In addition, a nested PCR protocol for detection of P. damselae based on 16S rRNA was developed. This PCR protocol was validated by testing 35 target and 24 nontarget pure cultures, and the detection limits obtained ranged from 1 pg to 10 fg of DNA (200 to 20 cells). A similar level of sensitivity was observed when the PCR protocol was applied to fish tissues spiked with bacteria. The PCR approach described in this paper allows detection of the pathogen in mixed plate cultures obtained from asymptomatic fish suspected to be carriers of P. damselae subsp. piscicida, in which growth of this bacterium cannot be visualized. Our results indicate that the selective primers which we designed represent a powerful tool for sensitive and specific detection of fish pasteurellosis.

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