Multi-genes Heredity Transformation And Extraneous Ssource Genes Expression In Populus

In this research, we obtained the transgenic insect-resistant poplar 741 through transferring agrobacterium rhizogenes Ri-plasmid T—DNA into transpolygenic strain 741, and preliminarily screened the pest-resisting and cutable transgenic aspen strain 741 through testing and analyzing the rooting characteristic, morphosis, content of Bt toxic protein and hormone in leaf; we also discussed the interaction of exogenous gene (BtCrylAc gene and Ri-plasmid T—DNA) expression in transpolygenic plant, as well as the affluence of environmental stress on exogenous gene expression. Furthermore, we transferred the rolB and rolC which were cloned from Ri-plasmid into common aspen 741 separately in order to study the functions of rolB and rolC and their appearances in transgenic plant, and screen the rootable transgenic strain. The main results are as follow:1. We preferred the transgenic insect-resistant poplar 741 aspen strain pb29 with high pest resistance and field seedling of transpolygenic 741 aspen strain (Bt toxic protein and Ri-plasmid T—DNA) as materials to study the growth characteristic of transpolygenic 741 aspen, and observe the shape and weight of leaf and length of leaf stalk. The results showed the leaves of transpolygenic 741 aspen were shorter and wider than that of without transferring agrobacterium rhizogenes, however the weight difference was not obviously, and the hormones were all increased above 2 times.2. We adopted the ELISA to test the Bt toxic protein in leaves from field transpolygenic aspen, and carded out the breeding experiment. The results showed the content of Bt toxic protein in transpolygenic aspen 741 leaf was lowed than that in double pest resisting aspen 741 (Pb29) without transferring agrobacterium rhizogenes; the fatality rate of Clostera anachoreta(Fabricius) bred with transpolygenic aspen 741 leaves was low, but the larval growth and development were inhibited obviously, so the normal life period was influenced; the fatality rate of Hvphantria cunea(Drurv) American white moth was still 100%, which showed the transgene of agrobacterium rhizogenes Ri-plasmid T—DNA inhibited the expression of Bt gene.3. We preferred the transgenic insect-resistant poplar 741 aspen strain pb29 and tissue seedling of transpolygenic 741 aspen strain as materials to study the responses of different transgenic strains on NaCl stress. The results showed after transferring the rol from Ri-plasmid T—DNA, the number of root system increased, length of root system reduced, contents of IAA and GA increased, and toxic protein expressing amount of BtCrylAc reduced; following the enhance of NaCl stress, the height of seedling, number of root system, contents of chlorophyll, IAA and GA reduced gradually, and the length of root system, Bt toxic protein expressing amount improved significantly, which showed the expressions of exogenous genes in transgenic aspen changed.4. We transferred rolB and rolC into 741 aspen separately by agrobacterium rhizogenes-mediated method, and obtained 15 cloned lines of trans-rolB and 2 cloned lines of trans-rolC; we detected rolB, rolC and NptⅡthrough PCR, which preliminarily proved that rolB and rolC had successfully integrated into the genome of 741 aspen.