Regeneration Of Populus Tomentosa Carr. Cv'BJHR01' And Research Of Bt Insect-resistant Genetic Engineering

Poplar is an important species of ecological shelter forest and industrial timber. But the poplar pests are so serious that enormous money loses every year. With the development of insect-resistant genetic engineering in poplar, some important insect-resistant genes, especilly Bt gene, have been successfully used for prevention and treatment of diseases, and we have achieved substantial results. The Populus tomentosa Carr. cv'BJHROl',which is superior varieties in North China, was used as the material in this study. The modified Bt886cry3Aa gene was taken as the target insect-resistant genes. We established the systems for tissue culture regeneration and genetic transformation, and finished the transformation of insect-resistant genes.Based on this, the new Bt protein expressed strain of Populus tomentosa was gained.Our detailed results are as following:(1)By exploring the media formulation, culture conditions and explant types, we established and optimized the tissue culture regeneration system of Populus tomentosa Carr. cv'BJHR01'.This is important to spread the fine varieties and preserve the germplasm resources. Moreover, our results set up the foundation for genetic engineering, and provides the scientific and technological support for the healthy development of the agroforestry ecosystem.(2) By testing the transient expression of gus gene, we establish and verify the feasibility of the entire genetic transformation system. And by groping the resistance to kanamycin of the transformants, we ascertain proper consistency of the antibiotic, and set the basis for the transformation of the insect-resistant gene.(3)We obtained the transgenetic Populus tomentosa Carr. cv'BJHROl'by agrobacterium-mediated genetic transformation, and there were 64 positive transgenic plants detected by PCR. PCR-Southern hybridization test validated the PCR results once again. The mBt-Cry3A ImmunoStrip could detect whether there is protein expression in insect-resistant plants, and the results showed that there were 12 positive reacted plants in the 64 PCR-positive plants. The mBt-Cry3A ELISA test re-verified the 12 positive plants which OD600 absorbance was generally higher than the control plants. All these proved that several insect-resistant transgenic poplar strains had been acquired.