Environmental Application of Nucleic Acid Hybridization

Sayler, Gary S.; Layton, Alice C.

doi:10.1146/annurev.mi.44.100190.003205

Cited by 174 publications

(51 citation statements)

References 67 publications

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“…Over the last decade nucleic acid hybridization has become a powerful tool for the detection of microorganisms (Saylor & Layton, 1990) and for the monitoring of population dynamics in microbial ecology (Stahl et al, 1988). Specific identification of individual microbial cells was recently achieved by hybridization with radioactively (Giovannoni et al, 1988) or fluorescently (DeLong et al, 1989; Amann et al, 1990a, b ; Tsien et al, 1990) labelled oligonucleotide probes directed against the naturally amplified, intracellular rRNA.…”

Section: -7028mentioning

confidence: 99%

Identification of single bacterial cells using digoxigenin-labelled, rRNA-targeted oligonucleotides

Zarda

Amann

Wallner

et al. 1991

Journal of General Microbiology

123

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Oligonucleotides were end-labelled with digoxigenin (DIG), chemically at the 5'-end or enzymically at the 3'-end. Following specific in situ hybridization of these probes to intracellular rRNA molecules, the hybrids were detected with anti-DIG Fab fragments labelled with fluorescent dyes. The antibody fragments penetrated through the bacterial cell periphery and specifically bound to their antigens. Probe-conferred and non-specific fluorescence per cell were quantified by flow cytometry and compared to values obtained with end-labelled fluorescent probes. The DIG reporter molecules could also be detected in whole fixed cells by antibodies labelled with either alkaline phosphatase or horseradish peroxidase. The penetration of the large antibody-enzyme complexes into the cells required lysozyme/EDTA treatment prior to the hybridization and has so far only been achieved for Gramnegative bacteria. This technique has the potential for significant signal amplification as compared to the fluorescently end-labelled oligonucleotides hitherto used for single cell identification in microbial ecology. Moreover, it can be used instead of fluorescent assays in natural samples showing autofluorescence.

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Section: -7028mentioning

confidence: 99%

Identification of single bacterial cells using digoxigenin-labelled, rRNA-targeted oligonucleotides

Zarda

Amann

Wallner

et al. 1991

Journal of General Microbiology

123

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“…Methodologies involving the detection of target nucleic acid sequences have been reviewed extensively (Hames & Higgins, 1985;Hazen & Jiminez, 1988;Sayler & Layton, 1990). Many applications of nucleic acid hybridization using environmental samples have involved probing immobilized DNA sequences fixed to nitrocellulose or nylon membranes.…”

Section: Detection Of Marker Genes Using Nucleic Acid Probesmentioning

confidence: 99%

Development of molecular methods for the detection of specific bacteria in the environment

Pickup¹

1991

Journal of General Microbiology

116

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“…In recent years, the use of molecular techniques to detect specific genes directly, especially in contaminated soil and sediment environments, has become more widespread. In particular, hybridization of catabolic gene probes and PCR amplification using specific primers have been used successfully to detect target genes in DNA extracted directly from the environment (Holben et al 1988, Sayler & Layton 1990, Walia et al 1990, Tsai & Olson 1991, Holben et al 1992, Herrick et al 1993, Joshi & Walia 1996, supplementing and sometimes circumventing the need for isolation and culturing of microorganisms. Of course, depen-dence upon cultures for probe development means that our environmental analyses are still somewhat constrained by the culture bias, but rapid progress has nevertheless been made with this approach.…”

Section: Introductionmentioning

confidence: 99%

Quantification of catechol 2,3-dioxygenase gene homology and benzoate utilization in intertidal sediments

Francis¹,

Francis²,

Golet³

et al. 1998

Aquat. Microb. Ecol.

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The catechol 2,3-dioxygenase (C230) gene, xylE, from the TOL plasmid pathway was used to probe naturally occurring bacterial communities in an intertidal microbial mat. Bound probe was quantified by densitometric analysis of slot blots using colorimetric detection of hybridization between the probe and total DNA extracts from the sediments. The C230 gene encodes the key ringbrealung enzyme of the toluene degradation pathway, of which benzoate is an intermediate. Radiotracer experiments using I4C-benzoate detected benzoate mineralization in these sedirnents, corroborating the presence of both the genetic potential and in situ actlvity for this transformation.

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Environmental Application of Nucleic Acid Hybridization

Cited by 174 publications

References 67 publications

Identification of single bacterial cells using digoxigenin-labelled, rRNA-targeted oligonucleotides

Identification of single bacterial cells using digoxigenin-labelled, rRNA-targeted oligonucleotides

Development of molecular methods for the detection of specific bacteria in the environment

Quantification of catechol 2,3-dioxygenase gene homology and benzoate utilization in intertidal sediments

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