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Detection of root mat associated Agrobacterium strains from plant material and other sample types by post‐enrichment TaqMan PCR
Author(s) -
Weller S.A.,
Stead D.E.
Publication year - 2002
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1046/j.1365-2672.2002.01506.x
Subject(s) - taqman , agrobacterium , biology , polymerase chain reaction , plasmid , primer (cosmetics) , pathogen , microbiology and biotechnology , biovar , transformation (genetics) , dna , bacteria , genetics , gene , chemistry , organic chemistry
Aims: The development of a fluorogenic, 5′ nuclease, TaqMan PCR assay for the detection of Ri‐plasmids from root mat inducing Agrobacterium biovar 1 strains. Methods and Results: A TaqMan probe and primer set were designed within the T‐DNA sequence of a known root mat inducing Agrobacterium strain. One hundred and ten Agrobacterium and closely related bacteria were tested using this novel PCR and compared with results from a conventional PCR which detects Ti and Ri‐plasmids. The Agrobacterium selective media, Medium 1A was modified into broth form for use as an enrichment of the pathogen from samples prior to the TaqMan PCR. Conclusions: The root mat pathogen was detected successfully from a range of sample types using the enriched fluorogenic PCR assay, negating the need for complex DNA extraction procedures and post‐PCR processing techniques such as gel electrophoresis. The technique is therefore a rapid and cost‐effective detection method. Significance and Impact of the Study: This is the first known report of a fluorogenic, 5′nuclease, TaqMan assay designed to detect an Agrobacterium plant pathogen. The method can be used as a model system for the detection of other Agrobacterium pathogens.