APPLICATION OF SUBTRACTION HYBRIDIZATION FOR THE DEVELOPMENT OF A RHIZOBIUM-LEGUMINOSARUM BIOVAR PHASEOLI AND RHIZOBIUM-TROPICI GROUP-SPECIFIC DNA-PROBE

W Streit, AJ Bjourson, JE Cooper, D Werner

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

A combined subtraction hybridization and polymerase chain reaction/amplification technique was used to develop a DNA probe which was specific for the Rhizobium leguminosarum biovar phaseoli and the Rhizobium tropici group. Total genomic DNA preparations from Rhizobium leguminosarum biovar viciae, Rhizobium leguminosarum biovar trifolii, Rhizobium sp., Agrobacterium tumefaciens, Rhizobium fredii, Bradyrhizobium japonicum, Bradyrhizobium ssp. and Rhizobium meliloti were pooled and used as subtracter DNA against total genomic DNA from the Rhizobium leguminosarum biovar phaseoli strain KIM5s. Only one round of subtraction hybridization at 65 degrees C was necessary to remove all cross-hybridizing sequences. Dot blot hybridizations with total genomic DNA of the eight subtracter organisms and 29 bacteria of different groups confirmed the high specificity of the isolated DNA sequences. Dot blot hybridizations with total genomic DNA from ten different R. leguminosarum biovar phaseoli and R. tropici strains resulted in strong hybridization signals for all strains tested. The DNA probe for the R. tropici and R. leguminosarum biovar phaseoli group was used for dot blot hybridization with DNA extracts from three tropical and one boreal soil. When correlated with data from Most Probable Number analyses the probe was capable of detecting as low as 3X10(4) homologous indigenous rhizobia per g soil. The technique offers great benefits for the development of DNA probes for monitoring bacterial populations in environmental samples.
LanguageEnglish
Pages59-67
JournalFEMS Microbiology Ecology
Volume13
Issue number1
Publication statusPublished - Oct 1993

Fingerprint

Rhizobium leguminosarum bv. phaseoli
Rhizobium tropici
DNA probes
hybridization
DNA
nucleic acid hybridization
genomics
Rhizobium
Sinorhizobium fredii
Rhizobium leguminosarum bv. viciae
Rhizobium leguminosarum bv. trifolii
Ensifer meliloti
Bradyrhizobium
Bradyrhizobium japonicum
Agrobacterium radiobacter
soil
polymerase chain reaction
nucleotide sequences
monitoring
bacteria

Cite this

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title = "APPLICATION OF SUBTRACTION HYBRIDIZATION FOR THE DEVELOPMENT OF A RHIZOBIUM-LEGUMINOSARUM BIOVAR PHASEOLI AND RHIZOBIUM-TROPICI GROUP-SPECIFIC DNA-PROBE",
abstract = "A combined subtraction hybridization and polymerase chain reaction/amplification technique was used to develop a DNA probe which was specific for the Rhizobium leguminosarum biovar phaseoli and the Rhizobium tropici group. Total genomic DNA preparations from Rhizobium leguminosarum biovar viciae, Rhizobium leguminosarum biovar trifolii, Rhizobium sp., Agrobacterium tumefaciens, Rhizobium fredii, Bradyrhizobium japonicum, Bradyrhizobium ssp. and Rhizobium meliloti were pooled and used as subtracter DNA against total genomic DNA from the Rhizobium leguminosarum biovar phaseoli strain KIM5s. Only one round of subtraction hybridization at 65 degrees C was necessary to remove all cross-hybridizing sequences. Dot blot hybridizations with total genomic DNA of the eight subtracter organisms and 29 bacteria of different groups confirmed the high specificity of the isolated DNA sequences. Dot blot hybridizations with total genomic DNA from ten different R. leguminosarum biovar phaseoli and R. tropici strains resulted in strong hybridization signals for all strains tested. The DNA probe for the R. tropici and R. leguminosarum biovar phaseoli group was used for dot blot hybridization with DNA extracts from three tropical and one boreal soil. When correlated with data from Most Probable Number analyses the probe was capable of detecting as low as 3X10(4) homologous indigenous rhizobia per g soil. The technique offers great benefits for the development of DNA probes for monitoring bacterial populations in environmental samples.",
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APPLICATION OF SUBTRACTION HYBRIDIZATION FOR THE DEVELOPMENT OF A RHIZOBIUM-LEGUMINOSARUM BIOVAR PHASEOLI AND RHIZOBIUM-TROPICI GROUP-SPECIFIC DNA-PROBE. / Streit, W; Bjourson, AJ; Cooper, JE; Werner, D.

In: FEMS Microbiology Ecology, Vol. 13, No. 1, 10.1993, p. 59-67.

Research output: Contribution to journalArticle

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