The Transformation And Expression Of Poplar 741 With Rol Gene

Ri plasmid contains rol genes which can induce formation of adventitious roots. The transformation of rol gene into plants can improve rooting ability of plants. However, some plants exhibited a series of variations, which influenced plant growth significantly. In order to study these variations, we optimize transformation systems of transfer rolB,rolC genes into poplar 741,and then get the transgenetic plants to identify the morphological character, the content of chloroplast, the character of fluorescence and the dynamic of endogenous hormones. Results were showed as follow:An efficient and stable system for poplar 741 mediated by agrobacterium rhizogenes with rolB,rolC gene was established through investigating the factors influencing the transformation efficiency. At last we obtain 8 stains of transgenetic rolB gene,and 18 stains of transgenetic rolC gene.The results showed that the optimization system is that adventitious bud differentiation of poplar 741 explants can reach to 97% with 6-BA 1.0 add NAA 0.1;the critical concentration of Km is 40 mg/L, the 400 mg/L Cef can efficacious control the Agrobacterium remain on poplar 741 explants as well as reduce the effect of CTX to adventitious bud differentiation; and the co-cultivation and liquid bacterium with AS 150 mg/L,10~15 minutes for infection, 3 days for co-culture, the conversion rate can be improved to 51%.The physiological indicators of transgenic plants showed that: Compared with poplar 741, most of transgenic plants with rolB gene increased root number and rooting rate, the high growth have no difference with poplar 741,chloroplast a,b and a+b content increased, and the chloroplast a,b and a+b have highly significant pozitive correlation. The Fv/Fm and PI value decreased,and chloroplast a,b,a+b and PI have negative correlation. There is a big change in hormone content, the IAA, ZT content increased, ABA content decreased, IAA/ABA value increased,which have consist with root number and rooting rate,while there is difference between IAA/ZT and the high growth. To transgenic plants with rolC gene, the plant increased root number,rooting rate and the highth become dwarf.Chloroplast a,b and a+b content decreased, and the chloroplast a,b and a+b have highly significant correlation. The Fv/Fm and PI value decreased,and chloroplast a,b,a+b and PI have significant pozitive correlation.The IAA,ABA,ZT content are all increased, IAA/ABA and IAA/ZT value are both increased,which have consiste with root number and high growth .Taking poplar 741 mediated by Agrobacterium rhizogenes with Ri plasmid as material, the result showed that the Agrobacterium on transgenic plant can be controlled basicly when the transgenic plants cultured in medium with 300 mg/L ceftiofur;the quantity and activity of the residual Agrobacterium on transgenic plant will be declined with the time prolong, and the Agrobacterium can not be cultured in YEB when the transgenic plant cultured in 18 month later. The residual Agrobacterium rhizogenes mainly distributed on transgenic plant surface and stem inside ,which seceriously effect the survival of transgenic plants and the PCR result .While the Agrobacterium on transgenic plant will be resume when the transgenic plant cultured in medium without ceftiofur;and the foreign Gene would flow once there are Agrobacterium rhizogenes residual on transgenic plants.