Effects Of Four Strains Of Erwinia Carotovora Subsp. Carotovora On The Activity Of Defense Enzymes Of Potato Tubers And The Cloning And Expression Of The Pathogenic Genes

Potato (Solanum tuberosum L.) is an important food crop. If there were encountered therain weather and unreasonably managed in the harvest and storage, pathogens maybe infectpotato tubers to cause rot, and impact the quality and price of potatoes. Therefore, the studyused four soft rot pathogenic strains (Ecc71、AC5070、AC5071、KD100) to infect the‘Atlantic’ virus-free potato tubers to determine the virulence of the pathogens and defenseenzyme activities of potato after inoculated, which aimed to know the infected ability of thesestrains and the response of defense enzymes in tubers. Then, the four pathogenic genes (pel、peh、cel、prt) form Ecc71strain were cloned and prokaryotic expression, which was designedto get the functional pathogenic genes and laid the foundation of further study pathogenesis.The results were showed as following:1. The ‘Atlantic’ virus-free potato tubers were infected by Ecc71、AC5070、AC5071andKD100strains, and the there were significant differences. In the whole infected process, thechange speed and degree of decay of tubers inoculated by KD100strain were greatest thanothers, but the responses of tubers inoculated by Ecc71were contrary. Meanwhile, the POD、PAL and PPO enzyme activities were higer than control (no inoculated) and the trends lastedincreasing until20h, then decreasing. But the trend of CAT enzyme activity of tubers afterinoculated was a constant declining trend, and the CAT enzyme activity of inoculated tuberswas lower than control.2. The test successfully cloned pel、peh、cel、prt genes sequences by identification ofdelay plasmid, PCR, digesting delay plasmid by restriction enzyme and sequencing. Then,analyzing the characterisitc of protein which the genes sequences were encoded, it showedthat Pel、Peh、Cel and Prt were stable protein. The conserved domain of Pel had pectate lyasecatalytic and virulence functions; the Peh had hydrolysis and virulence function; the Cel hadcellulose binding domain and carbohydrate binding domain, which catalyzed glycosidehydrolysis; the Prt had a function of catalyzing the metallopeptidases.3. The proteins encoded by pel、peh、cel、prt genes induced by IPTG were efficientexpressed, and the protein molecular weight were37.23KDa,42KDa,48.29KDa and38.83KDa, respectively, which were consistent with the size of the expected. Among these proteins, the expression weight of Peh was lower than others. In addition, the enzyme activties weresignificantly increased than no induced by43.6%,75%,51.6%and65.3%, respectively, andthe increased rage of Peh enzyme activity were the highest than no induced. The above resultsindicated that the functional genes were successfully cloned.