An ABC Transporter Protein and Molecular Diagnoses of Xanthomonas arboricola pv. pruni Causing Bacterial Spot of Stone Fruits

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dc.contributor.advisor Turner Sutton, Committee Member en_US
dc.contributor.advisor Eric Davis, Committee Member en_US
dc.contributor.advisor David Ritchie, Committee Chair en_US
dc.contributor.advisor Peter Lindgren, Committee Member en_US Pagani, Maria Cristina en_US 2010-04-02T18:55:47Z 2010-04-02T18:55:47Z 2005-02-27 en_US
dc.identifier.other etd-10042004-232356 en_US
dc.description.abstract The purpose of the research was to develop a detection and identification system for Xanthomonas arboricola pv. pruni using polymerase chain reaction (PCR) analysis and Southern hybridization with a DNA probe. Random amplified polymorphic DNA (RAPD) analysis was used to identify a specific DNA sequence strictly associated and conserved among all X. arboricola pv. pruni strains tested and obtained from various locations and hosts. PCR primers Y17CoF and Y17CoR specific to this DNA fragment were synthesized and evaluated as a diagnostic tool. Primers amplified a 943-bp DNA fragment in all strains previously identified as X. arboricola pv. pruni on the basis of biochemical and physiological tests, and failed to amplify DNA from other xanthomonads and non-xanthomonads including saprophytes and epiphytes associated with Prunus spp. The PCR assay detected between 25 and 50 cells. A digoxigenin-labeled DNA probe, XPRUNI14, was developed and used to detect an extensive collection of this bacterium through dot-blot and Southern analysis. Results indicated that X. arboricola pv. pruni could be accurately detected and identified by PCR analysis and probe hybridizations on symptomatic and asymptomatic plant materials avoiding the need for prior isolation of this phytopathogen. Sequence analysis of this 943-bp DNA fragment revealed the presence of an open reading frame (ORF) predicted to encode a putative protein of 243 amino acids with a calculated molecular mass of 26.8 kDa. In silico analysis predicted this protein to share similarity to the ABC transporter family, with a FtsX conserved domain possibly involved in cell division. In the present study we overexpressed the putative protein and results were confirmed with western and immunoblot analysis. Gene disruption with a streptomycin cassette resulted in small colony phenotype, with no evidence of bacterial mass increase suggestive of impaired division that led them to cell death. en_US
dc.rights I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to NC State University or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. en_US
dc.subject bacteria en_US
dc.subject detection en_US
dc.subject diagnosis en_US
dc.subject pruni en_US
dc.subject arboricola en_US
dc.subject bacterial spot en_US
dc.subject stone fruits en_US
dc.subject ABC protein en_US
dc.subject Xanthomonas en_US
dc.title An ABC Transporter Protein and Molecular Diagnoses of Xanthomonas arboricola pv. pruni Causing Bacterial Spot of Stone Fruits en_US PhD en_US dissertation en_US Plant Pathology en_US

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