Preparation Of A Monoclonal Antibody And Construction Of Single ChainVariable Fragment (scFv) Antibody Library Against HrpA Protein Of Pseudomonas Syringae

In plant pathogenic bacteria, there is a gene family that regulates the bacteria triggering hypersensitive reaction and pathogenicity (hrp) in plant. In these genes. hrpA encodes a protein named as HrpA. HrpA is a major component of the filamentous surface appendage, Hrp pilus, which plays a very important role in the interaction between bacteria and their host plant. It has been demonstrated that there is a direct and decisive relationship between existence of the pilus and its causing Hrp protein in host, giving us hints of the possibility to improve plant resistance by blocking assembly of the Hrp pilus through expression of engineered antibodies in plant.In this study, the HrpA protein of Pseudomonas syringae pv. tomato DC3000 was expressed in Escherichia coli, purified as the antigen to immunise BLAB/c mice to preparate Monoclonal Antibody.The scFv DNA was amplified by PCR with cDNA reverse-transcribed from extracted RNA of hybidoma cells against HrpA protein and ligated into a phagemid vector pCANTAB-5E, and the ligated product was then transformed into E. coli TG1 to yield recombinant phages after infection with helper phage M13KO7.In a summary, our results will provide new gene resources for gene engineering for plant resistance and plant antibody.