PCR amplification of 16S rDNA from lyophilized cell cultures facilitates studies in molecular systematics

Wisotzkey, Jeffrey D.; Jurtshuk, Peter; Fox, George E.

doi:10.1007/bf02092099

Cited by 65 publications

(38 citation statements)

References 6 publications

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“…DNA from Prosthecobacter was isolated according to the method of Wisotzkey et al (63). DNA from A. muciniphila, C. flavus, O. terrae, and isolate Ellin514 was extracted using a modified protocol to inhibit high nuclease activity.…”

Section: Methodsmentioning

confidence: 99%

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Characterization and Evolution of Cell Division and Cell Wall Synthesis Genes in the Bacterial Phyla Verrucomicrobia , Lentisphaerae , Chlamydiae , and Planctomycetes and Phylogenetic Comparison with rRNA Genes

et al. 2008

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In the past, studies on the relationships of the bacterial phyla Planctomycetes, Chlamydiae, Lentisphaerae, and Verrucomicrobia using different phylogenetic markers have been controversial. Investigations based on 16S rRNA sequence analyses suggested a relationship of the four phyla, showing the branching order Planctomycetes, Chlamydiae, Verrucomicrobia/Lentisphaerae. Phylogenetic analyses of 23S rRNA genes in this study also support a monophyletic grouping and their branching order-this grouping is significant for understanding cell division, since the major bacterial cell division protein FtsZ is absent from members of two of the phyla Chlamydiae and Planctomycetes. In Verrucomicrobia, knowledge about cell division is mainly restricted to the recent report of ftsZ in the closely related genera Prosthecobacter and Verrucomicrobium. In this study, genes of the conserved division and cell wall (dcw) cluster (ddl, ftsQ, ftsA, and ftsZ) were characterized in all verrucomicrobial subdivisions (1 to 4) with cultivable representatives (1 to 4). Sequence analyses and transcriptional analyses in Verrucomicrobia and genome data analyses in Lentisphaerae suggested that cell division is based on FtsZ in all verrucomicrobial subdivisions and possibly also in the sister phylum Lentisphaerae. Comprehensive sequence analyses of available genome data for representatives of Verrucomicrobia, Lentisphaerae, Chlamydiae, and Planctomycetes strongly indicate that their last common ancestor possessed a conserved, ancestral type of dcw gene cluster and an FtsZ-based cell division mechanism. This implies that Planctomycetes and Chlamydiae may have shifted independently to a non-FtsZ-based cell division mechanism after their separate branchings from their last common ancestor with Verrucomicrobia.

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Section: Methodsmentioning

confidence: 99%

“…The mixture was incubated for 1 h at 55°C. The subsequent steps were as described by Wisotzkey et al (63).…”

Section: Methodsmentioning

confidence: 99%

Characterization and Evolution of Cell Division and Cell Wall Synthesis Genes in the Bacterial Phyla Verrucomicrobia , Lentisphaerae , Chlamydiae , and Planctomycetes and Phylogenetic Comparison with rRNA Genes

et al. 2008

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“…B. psychrophilus W16AT and W5 and B. globisporus W25T chromosomal DNAs were prepared and polymerase chain reaction (PCR) gene amplifications were carried out as previously described (28). The two primers used were complementary to phylogenetically conserved portions of the 5' and 3' ends of the 16s rRNAs and contained 5' restriction site polylinker regions to facilitate cloning.…”

Section: Methodsmentioning

confidence: 99%

How Close Is Close: 16S rRNA Sequence Identity May Not Be Sufficient To Guarantee Species Identity

Fox

Wisotzkey

Jurtshuk

1992

International Journal of Systematic Bacteriology

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1,174

731

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16s rRNA (genes coding for rRNA) sequence comparisons were conducted with the following three psychrophilic strains: Bacillus globisporus W25* (T = type strain) and Bacillus psychrophilus W16AT, and W5.These strains exhibited more than 99.5 % sequence identity and within experimental uncertainty could be regarded as identical. Their close taxonomic relationship was further documented by phenotypic similarities. In contrast, previously published DNA-DNA hybridization results have convincingly established that these strains do not belong to the same species if current standards are used. These results emphasize the important point that effective identity of 16s rRNA sequences is not necessarily a sufficient criterion to guarantee species identity. Thus, although 16s rRNA sequences can be used routinely to distinguish and establish relationships between genera and well-resolved species, very recently diverged species may not be recognizable.

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“…For DNA extraction according to method I, bacteria were resuspended from the PC filter pieces by vortexing with 2 ml of natural mineral water and pelleted by centrifugation (14,000 rpm, 10 min). DNA from pelleted bacteria was extracted by enzymatic cell lysis and chloroform treatment according to a previously established protocol (55). Method II involved the resuspension of PC filter pieces in 400 l of TE buffer (10 mM Tris-HCl, 1 mM EDTA [pH 8.0]).…”

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confidence: 99%

Diversity of Bacteria Growing in Natural Mineral Water after Bottling

Loy

Beisker²,

Meier

2005

Appl Environ Microbiol

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Bacterial growth occurs in noncarbonated natural mineral waters a few days after filling and storage at room temperature, a phenomenon known for more than 40 years. Using the full-cycle rRNA approach, we monitored the development of the planktonic bacterial community in a noncarbonated natural mineral water after bottling. Seven 16S rRNA gene libraries, comprising 108 clones in total, were constructed from water samples taken at various days after bottling and from two different bottle sizes. Sequence analyses identified 11 operational taxonomic units (OTUs), all but one affiliated with the betaproteobacterial order Burkholderiales (6 OTUs) or the class Alphaproteobacteria (4 OTUs). Fluorescence in situ hybridization (FISH) was applied in combination with DAPI (4,6-diamidino-2-phenylindole) staining, viability staining, and microscopic counting to quantitatively monitor changes in bacterial community composition. A growth curve similar to that of a bacterium grown in a batch culture was recorded. In contrast to the current perception that Gammaproteobacteria are the most important bacterial components of natural mineral water in bottles, Betaproteobacteria dominated the growing bacterial community and accounted for 80 to 98% of all bacteria detected by FISH in the late-exponential and stationary-growth phases. Using previously published and newly designed genusspecific probes, members of the betaproteobacterial genera Hydrogenophaga, Aquabacterium, and Polaromonas were found to constitute a significant proportion of the bacterial flora (21 to 86% of all bacteria detected by FISH). For the first time, key genera responsible for bacterial growth in a natural mineral water were identified by applying molecular cultivation-independent techniques.

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PCR amplification of 16S rDNA from lyophilized cell cultures facilitates studies in molecular systematics

Cited by 65 publications

References 6 publications

Characterization and Evolution of Cell Division and Cell Wall Synthesis Genes in the Bacterial Phyla Verrucomicrobia , Lentisphaerae , Chlamydiae , and Planctomycetes and Phylogenetic Comparison with rRNA Genes

Characterization and Evolution of Cell Division and Cell Wall Synthesis Genes in the Bacterial Phyla Verrucomicrobia , Lentisphaerae , Chlamydiae , and Planctomycetes and Phylogenetic Comparison with rRNA Genes

How Close Is Close: 16S rRNA Sequence Identity May Not Be Sufficient To Guarantee Species Identity

Diversity of Bacteria Growing in Natural Mineral Water after Bottling

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