Heteroduplex structures in 16S-23S rRNA intergenic transcribed spacer PCR products reveal ribosomal interoperonic polymorphisms within single<i>Frankia</i>strains

Gtari, Maher; Brusetti, Lorenzo; Chérif, Ameur; Boudabous, Abdellatif; Daffonchio, Daniele

doi:10.1111/j.1365-2672.2007.03329.x

Cited by 5 publications

(4 citation statements)

References 37 publications

(93 reference statements)

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“…The presence of these easily distinguished, slower-migrating forms is commonly exploited for heteroduplex analysis of microsatellite variability (Barros et al, 2005; Gtari et al, 2007). We then performed heteroduplex analysis by PCR amplification of the luciferase constructs containing 2, 4, 5, and 6 GGGGC-repeats (Section 2.5 ) as templates, including a construct that was engineered to contain a (GGGGC) 3 repeat (Section 2.7 ).…”

Section: Resultsmentioning

confidence: 99%

“…Genotypes containing SNPs were amplified at least two times for confirmatory sequencing. All genotypes were verified with respect to the number of (GGGGC) n repeats by performing a separate PCR amplification using the AhRgel-F and AhRgel-R primers that span −57 to +8 nt relative to the TSS and subsequent heteroduplex analysis (Barros et al, 2005; Gtari et al, 2007) of products separated by polyacrylamide gel electrophoresis (PAGE). Differing concentrations of DNA and MgCl 2 were used in this second amplification to ensure that both alleles were amplified.…”

Section: Methodsmentioning

confidence: 99%

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Analysis of the AHR gene proximal promoter GGGGC-repeat polymorphism in lung, breast, and colon cancer

Spink

Bloom

et al. 2015

Toxicology and Applied Pharmacology

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The aryl hydrocarbon receptor (AhR) regulates expression of numerous genes, including those of the CYP1 gene family. With the goal of determining factors that control AHR gene expression, our studies are focused on the role of the short tandem repeat polymorphism, (GGGGC)n, located in the proximal promoter of the human AHR gene. When luciferase constructs containing varying GGGGC repeats were transfected into cancer cell lines derived from lung, colon, and breast, the number of GGGGC repeats affected AHR promoter activity. The number of GGGGC repeats was determined in DNA from 327 humans and from 38 samples representing 5 species of non-human primates. In chimpanzees and 3 species of macaques, only (GGGGC)2 alleles were observed; however, in western gorilla, (GGGGC)n alleles with n = 2, 4, 5, 6, 7, and 8 were identified. In all human populations examined, the frequency of (GGGGC)n was n = 4>5≫2, 6. When frequencies of the (GGGGC)n alleles in DNA from patients with lung, colon, or breast cancer were evaluated, the occurrence of (GGGGC)2 was found to be 8-fold more frequent among lung cancer patients in comparison with its incidence in the general population, as represented by New York State neonates. Analysis of matched tumor and non-tumor DNA samples from the same individuals provided no evidence of microsatellite instability. These studies indicate that the (GGGGC)n short tandem repeats are inherited, and that the (GGGGC)2 allele in the AHR proximal promoter region should be further investigated with regard to its potential association with lung cancer susceptibility.

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Analysis of the AHR gene proximal promoter GGGGC-repeat polymorphism in lung, breast, and colon cancer

Spink

Bloom

et al. 2015

Toxicology and Applied Pharmacology

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“…Some genes and operon organisation show some distinctiveness for each of the four clusters ( Supplementary Figure 1 ). There are two ribosomal operons for clusters 1 and 2 strains and three for clusters 3 and 4 ( Gtari et al, 2007 ). Nitrogenase complexes encoding ( nif ) genes are totally absent in the asymbiotic cluster 4 (not retrieved in the draft genome sequence, except for F. asymbiotica strain M16386 T ).…”

Section: Resultsmentioning

confidence: 99%

Taxogenomic status of phylogenetically distant Frankia clusters warrants their elevation to the rank of genus: A description of Protofrankia gen. nov., Parafrankia gen. nov., and Pseudofrankia gen. nov. as three novel genera within the family Frankiaceae

Gtari

2022

Front. Microbiol.

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The genus Frankia is at present the sole genus in the family Frankiaceae and encompasses filamentous, sporangia-forming actinomycetes principally isolated from root nodules of taxonomically disparate dicotyledonous hosts named actinorhizal plants. Multiple independent phylogenetic analyses agree with the division of the genus Frankia into four well-supported clusters. Within these clusters, Frankia strains are well defined based on host infectivity range, mode of infection, morphology, and their behaviour in culture. In this study, phylogenomics, overall genome related indices (OGRI), together with available data sets for phenotypic and host-plant ranges available for the type strains of Frankia species, were considered. The robustness and the deep radiation observed in Frankia at the subgeneric level, fulfilling the primary principle of phylogenetic systematics, were strengthened by establishing genome criteria for new genus demarcation boundaries. Therefore, the taxonomic elevation of the Frankia clusters to the rank of the genus is proposed. The genus Frankia should be revised to encompass cluster 1 species only and three novel genera, Protofrankia gen. nov., Parafrankia gen. nov., and Pseudofrankia gen. nov., are proposed to accommodate clusters 2, 3, and 4 species, respectively. New combinations for validly named species are also provided.

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“…could be used for in situ measurement of nitrite-oxidizing activity to improve implementation of traditional nitrification (17). Furthermore, pre-16S 3Ј rRNA sequence information has been very important in taxonomic and ecological studies (16) and has been used to differentiate closely related strains in a broad diversity of bacterial genera and species, such as Sphingomonas (37), Frankia (15), and Pseudomonas avellanae (25).…”

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Reverse Transcription of 16S rRNA To Monitor Ribosome-Synthesizing Bacterial Populations in the Environment

Stroot

Oerther

2009

Appl Environ Microbiol

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Identification and quantification of phylogenetically defined bacterial populations in the environment are often performed using molecular tools targeting 16S rRNA. Fluorescence in situ hybridization has been used to monitor the expression and processing of rRNA by targeting the 3 tail of precursor 16S rRNA. To expand this approach, we employed reverse transcription of total RNA using primer S-D-Bact-0338-a-A-18. Length heterogeneity detected by slab gel analysis, denaturing high-performance liquid chromatography (DHPLC) was used to differentiate the 5 tail of the precursor from mature 16S rRNA, and the relative abundance of the precursor compared to the abundance of mature 16S rRNA was shown to be a sensitive indicator of the physiologic state of Acinetobacter calcoaceticus ATCC 23055T . Our results demonstrate that this is a sensitive and reliable method with a detection limit of 10 ng of single-stranded DNA. The assay was also used to differentiate among precursor 16S rRNA levels with mixed pure cultures, as well as to examine the response of a mixed activated sludge culture exposed to fresh growth medium and the antibiotic chloramphenicol. The results of this study demonstrate that this assay is a novel reverse transcription assay that simultaneously measures the mature and precursor 16S rRNA pools for mixed bacterial populations in an engineered environment. Furthermore, collection of the reverse transcription products derived from activated sludge samples by the DHPLC approach enabled identification of the active bacterial genera. Comparison of 16S and precursor 16S rRNA clone library results indicated that the precursor 16S rRNA library is a more sensitive indicator for active bacteria in engineered environmental samples.For nearly 20 years, the "full-cycle 16S rRNA approach" has been employed to identify, enumerate, and determine the spatial organization of bacterial populations in environmental samples without the need for cultivation (1). The results of these studies have profoundly impacted the view of microbial diversity as a regulator of the global biosphere (10). During the development, demonstration, and maturation of the use of 16S rRNA-targeted molecular biology tools for bacterial identification, a number of researchers expanded the value of these methods in an attempt to determine simultaneously the identity and physiological status of bacterial populations. For example, Poulsen and coworkers quantified the intensity of the fluorescent signal from whole-cell fluorescence in situ hybridizations (FISH), targeting 16S rRNA as a predictor of ribosome abundance in bacterial cells of young and mature biofilms (31).The correlation between the cellular ribosome (rRNA) content and the growth rate was one of the earliest and most fundamental observations in microbial physiology (31). An approximately 10-fold increase in the ribosome level is observed when the doubling time of Escherichia coli decreases from 100 min to 24 min. During rapid growth (doubling time, Ͻ1 h), over 50% of the total RNA produced ...

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Heteroduplex structures in 16S-23S rRNA intergenic transcribed spacer PCR products reveal ribosomal interoperonic polymorphisms within singleFrankiastrains

Cited by 5 publications

References 37 publications

Analysis of the AHR gene proximal promoter GGGGC-repeat polymorphism in lung, breast, and colon cancer

Analysis of the AHR gene proximal promoter GGGGC-repeat polymorphism in lung, breast, and colon cancer

Taxogenomic status of phylogenetically distant Frankia clusters warrants their elevation to the rank of genus: A description of Protofrankia gen. nov., Parafrankia gen. nov., and Pseudofrankia gen. nov. as three novel genera within the family Frankiaceae

Reverse Transcription of 16S rRNA To Monitor Ribosome-Synthesizing Bacterial Populations in the Environment

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