Cloning And Expression Of Harpin From Erwinia Carotovora Subsp. Carotovora

Harpin encoded by hrpN gene in Erwinia carotovora subsp, carotovoracan induce hypersensitive reaction and systemic acquired resistance onnon-host plants.In this research, the hrpNEcc9 gene was amplified by PCR from the strainSE9 (Erwinia carotovora subsp, carotovora) total DNA. After sequencingand analyzing the gene hrpNEcc9 by corresponding software, the resultindicated that HarpinEcc9 encoded by hrpNEcc9 was a heat-stable,glycine-rich, cysteine-free protein, and hrpNEcc9 had higher similarities toreported hrp gene from other Erwinia carotovora, which is useful toresearching the relationship of structure and function.Then, the target gene was cloned into the secretion-expression vectorpExSecâ… to obtain pES-hrpN. The recombinant plasmids were transformedinto E.coli BL21(DE3). After inducing the recombinant bacteria with IPTG,the result showed that the fusion protein SZZ-Harpin not only secreted andexpressed via typeâ…¡secretion system, but also introduced HR.In order to expressing Harpin without inducement, the gene that encodesSZZ-Harpin was subcloned into the consititutive-expression vectorpCPP2139. The recombinant plasmid pCS-hrpN was transformed into E.coliDH5a.The fusion protein SZZ-Harpin expressed but exited in deposit. ByHR test, the protein and the recombinant bacteria couldn't induce HR, socouldn't introduce plant resistance. Therefore, future work is researching andconstructing appropriate consititutive-expression plasmid for biocontrolwork.