Molecular detection of a bacterial contaminant Bacillus pumilus in symptomless potato plant tissue cultures |
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| Authors: | Isenegger D A Taylor P W J Mullins K McGregor G R Barlass M Hutchinson J F |
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| Institution: | Institute for Horticultural Development, Agriculture Victoria, Knoxfield, Private Bag 15, 3180, Ferntree Gully Delivery Centre, Victoria, Australia. daniel.isenegger@nre.vic.gov.au |
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| Abstract: | An aberrant random amplified polymorphic DNA (RAPD) marker in genomic DNA of tissue culture plantlets was frequently observed during a comparison of DNA fingerprints derived from potato germplasm grown in tissue culture and the field. The RAPD marker was cloned, sequenced and determined to be of bacterial origin. A bacterial contaminant was isolated from the tissue culture plants and identified as a Bacillus pumilus. A set of sequence characterised amplified region (SCAR) primers were designed from the sequence of the cloned fragment and tested for the specific detection of B. pumilus. Polymerase chain reaction-restriction fragment length polymorphisms (PCR-RFLPs) were also used to generate B. pumilus profiles specific to our isolate in order to test and confirm the sequence homology of amplified markers generated from a range of DNA samples isolated from tissue culture plants and pure isolates of B. pumilus-like bacteria. |
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