Establishment And Research A Genetic Transformation System Of Macleaya Cordata

Macleaya cordata is an important and secure medicinal herb which has a wide market prospect. Genetic engineering is a new method for genetic improvement of, and can solve the problem of shrinking resources and supply shortage of Macleaya cordata. Our had been to The expression vector of pCAMBIA1300-MtDREB1C has been constructed by our laboratory and transferred into Agrobacterium. Through the process of infection blades of Macleaya cordata by Agrobacterium, the Callus was produced and the regeneration was accomplished. The relative influence factors of Agrobacterium mediated method were optimized preliminary, and a viable and effective genetic transformation system of macleaya cordata was established. The experiment results provided adequate material for the study of gene function for MtDREB1C.1. Leaf and stem of Macleaya cordata can produce callus after induced by kanamycin. It was shown that the leaf of Macleaya cordata was more suitable than stem section as an acceptor material. The higher concentration of kanamycin, the higher mortality of explants, and more callus would brown and dehydration. The optimum concentration range of kanamycin was determined as 25?50 mg/L by the induction experiment of leaf callus with kanamycin.2.Leaf can produce callus by the induction with cefotaxime or timentin. The results showed that cefotaxime is more suitable than timentin as an Agrobacterium inhibitor. The optimum concentration range of cefotaxime was selected as 300?400 mg/L to ensure the normal development of explants and prevent the Agrobacterium overgrowth,3. The leaves of Macleaya cordata which infected by Agrobacterium were transferred to fresh primary callus induction medium containing 50 mg/L kanamycin and 300 mg/L cefotaxime. After repeated screening, positive cells could regenerate transformed plants by the form of embryogenesis, and the regeneration rate of callus was 16.7%.4. By PCR and RT-PCR detection, the results showed that the corresponding strips of MtDREB1C gene could be amplified by the positive transformed plants only. The research was preliminary confirmed that MtDREB1C gene was successfully integrated into the genome of Macleaya cordata, and the emergence rate of positive seedlings of Macleaya cordata was 6%.