CLONING OF PLANT DEFENSE GENES PvPGIP2 AND TaLTP4 AND THEIR TRANSFORMATION TO WHEAT

Polygalacturcuase-inhibing protein(PGIP) and Lipid-transfer protein(LTP) were defense proteins produced in plant,which inhibit the growth of pathogenic fungi.In this study,a PGIP encoding gene in Phaseouls vulgaris(PvPGIP2) and a LTP encoding gene in Triticum aestivum(TaLTP4) were successfully cloned by using homologous gene cloning strategy and RT-PCR technique.By using pAHC25 as a frame vector,we constructed PvPGIP2and TaLTP4binary expression vector.Then the binary expression vector was through preliminary screening by colony PCR and restriction enzyme digest identification and sequencing analysis.The sequencing result showed that the PvPGIP2and TaLTP4binary expression vector was successfully constructed.In this transformation vector,the PvPGIP2and TaLTP4genes were respectively driven by the maize ubiquitin(Ubi) promoter and terminated by the 3′non-transcribed region of the Agrobacterium tumefaciensnopaline synthase gene(Tnos).Through microprojectile-mediated bombardment,the binary expression vector was transformed to 2,000 mature callus of wheat variety Yangmai 18.After twice rounds of Bialaphos screening 112 regeneration plants were finally obtained.The living transformed plants were subjected to PCR detection using primers specific to expression vector gene to conduct,12 positive plants with PvPGIP2and TaLTP4were obtained,and the conversion rate was 0.6%.