| Maize is the world’s important food, forage crop and industrial processing raw material. It can bring high economic benefits. Maize sheath blight is one of the main diseases of maize seriously affecting the production of maize, its pathogenic bacteria is Rhizoctonia solani Kuhn. The traditional chemical and agricultural control are still the main methods to prevent and control the sheath blight of maize. However, these prevention methods are time-consuming and with low effiency. In order to generate resistant maize to sheath blight, we cloned transcription factor Pti4gene from tomato and antibacterial peptide Rs-AFP2gene from radish seeds and transferred them into maize by Agrobacterium-mediated transformation.The cloning strategy is to construct a binary expression vector containing transcription factor Pti4gene and antimicrobial peptide Rs-AFP2gene. These two genes have their own promoter and terminator, respectively. The Pti4gene was inserted into the multiple cloniong site of pCAMBIA3301and was controlled by a promoter UBI-1, whereas, the Rs-AFP2was controlled by promoter CaMV35S. The created binary vector was designed as pCA3301-Pti4-Rs. This vector was verified by enzyme digestion, PCR detection and sequencing. The results showed that each fragment was linked into the binary expression vector correctly.The bivalent expression vector was used for gene engineering mediated by Agrobacuterium-mediated transformation. Maize varieties A188and Hi II were the target materials in the gene transformation experiments. There were47transgenic maize plants were obtained. Nested PCR analyses showed that15transgenic maize plants appeared PCR positive. Painting0.01%Basta on seedling leaf of transgenic plant, there was no necrosis. We tested disease resistance of the transgenic maize plants. Leaves from15transgenic plants were inoculated with R. solani Kuhn. We found that the transgenic lines were resistant to the R. solani Kuhn after48h. |
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