Complementation of an Erwinia carotovora subsp. carotovora protease mutant with a protease-encoding cosmid |
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Authors: | Caitilyn Allen Verlyn K. Stromberg Franzine D. Smith George H. Lacy Mark S. Mount |
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Affiliation: | (1) Department of Plant Pathology, Physiology, and Weed Science, Virginia Polytechnic Institute and State University, 24061 Blacksburg, VA, USA;(2) Department of Plant Pathology, University of Massachusetts, 01003 Amherst, MA, USA |
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Abstract: | Summary We report the complementation of a genetic lesion in the genome of Erwinia carotovora subsp. carotovora (Ecc), a pathogenic bacterium that incites soft rot of plants. A Sau3AI genomic library of Ecc was constructed using the conjugal cosmid pLAFR-3 as a vector. Sixteen cosmid clones encoding various plant tissue-degrading enzymes were identified, including a proteolytic clone, five cellulolytic clones, and ten pectolytic clones. We detected a mutant of Ecc with no proteolytic activity following transposon mutagenesis with an unstable Tn5-carrying plasmid. Conjugal transfer of the protease-encoding cosmid to this mutant restored near-wildtype extracellular protease production. Further manipulation and study of genes encoding pathogenic determinants in Ecc will be possible using this system. |
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Keywords: | Transposon mutagenesis Soft rot Phytopathogenicity |
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