| Most of plant pathogenic bacteria have the hrp-genes(hypersensitive response and pathogenicity genes), such as Pseudomonas, Erwinia, Xanthomonas and Ralstonia. The hrp cluster is not only responsible for causing obvious macroscopic symptoms in a susceptible host plant, but also they elicit the hypersensitive reaction (HR) in resistant host or in nonhost plants. When the pathogenic bacteria invade into the plant cells, proteins which are encoded by the hrp-genes assemble to a cell surface appendage, called Hrp pilus. The Hrp pilus serves as a conduit for the transfer of bacterial effector proteins into the plant cell cytosol. It plays an important role in the plant-pathogenic bacteria system.The sequence of P. syringae pv. tomato DC3000 HrpA gene was found in NCBI, and the expression vector pET32a(+)-HrpA was constructed . Then the recombinant HrpA protein was successfully expressed in Escherichia coli BL21. In order to assay antibody activity in sera without thioredoxin (Trx)'s interference, we also cloned HrpA gene into expression vector pET28a(+)and successfully purified the recombinant protein as assay protein.The BALB/c mice(6-8 weeks old) were immunized by the HrpA protein expressed by vector pET32a(+)-HrpA, then the serum titer were tested by ELISA. After total RNA was extracted from the spleen, cDNA was obteined by RT-PCR. The VH and VL genes were amplified separately, then the scFv genes was assembled with a Linker. The assembled scFv genes were cloned into a phage vector pCANTAB-5E to establish the primary phage library. Display antibody which contain its antigen-binding capability were enriched by panning process. With this method, a phage clone scFv(SF1) which showing the strongest antigen-binding capability was obtained after six rounds panning, then infected the E.coli HB2151 to achieve soluble expression of the scFv. It provided good resource for further research on transgenic plant and plant antibody.
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