关键词: 柑橘, 溃疡病, pthA, 抗血清, 病害抑制

Abstract:

The sequence encoding three nuclear localizing signals (NLSs) at the C-terminal of pthA, was amplified by PCR from the plasmid of Xanthomonas axonopodis pv. citri. and cloned into PET32a(+) vector. The recombinant plasmid named PthA-NLS was identified by restriction digestiion and sequence analysis. The sequence of the cloned NLSs had 99.9% of similarity with that of pthA in GenBank. The recombinant fusion protein was expressed in E.coli BL21 (DE3) and analyzed by 12% SDS-PAGE. A 48 kD of recombinant fusion protein was purified with Ni2+-NTA resin. The antiserum was obtained by immunizing Balb/c mouse with the purified recombinant protein and then identified by Western Blotting and ELISA analysis. The result demonstrated that the antiserum could specifically bind to the recombinant protein and the PthA from X. axonopodis pv. citri., as well as to partially inhibit the disease development. The successful cloning, expression and the preparation of PthA-NLS specific mouse antiserum offered the ways for better understanding the pathogenesis of pthA and the development of a rapid method for molecular detection of citrus canker.

Key words: Citrus, bacterial canker disease, pthA, antiserum, disease inhibition