High Efficient System Establishment On Plant Regeneration And Study On Genetics Transformation In Puna Chicory (Cichorium Intybus L.)

Puna Chicory (Cichorium Intybus L.), a perennial herbaceous plant of Compositae family, is aboriginal in Europe. It is made use of for the material of feedstuff and sugar refining. Puna Chicory is with high adaptability, with rich root ,with high production, with rich nourishment that the raw protein is about from 16.44% to 27.35%. It is proved a dainty pasture by feeding it to the cattles and the birds. It was migrated from New Zealand to our country. It is popular with herdsman because of its adaptability, dainty, high capacity of resistance to insect. Puna Chicory is a kind of pasture that has a promotional value. At present, the research about Puna Chicory is most on the culture technology for high yield. But the study on tissue culture and gene transformation is little.The leaves from the experiment field are the material. After disinfection with 0.1% HgCl2, the leaves were inoculated to the MS culture medium with different phytohormone combination to induce the callus and to differentiate the buds. We analyzed the effect of different phytohormone combination and different phytohormone concentration on the inducement of callus, buds and root. The result showed that 6-BA+IBA is the ideal phytohormone combination to induce the callus and to differentiate the buds. The culture medium MS+6-BA 2.0mg/L+IBA 0.1mg/L is suitable for the inducement of callus and the differentiation of buds, because the speed of callus inducement in this culture medium is faster than in other culture medium. And the number of buds differentiated in the single explant in this culture medium is more than in other culture medium. During the course of the root inducement, the culture medium 1/2MS is better than other culture medium. Active carbon was proved no good to the regeneration of root.Plant expression vector pBI121-gfp was constructed based on the vector pBI121. The vector pBI121 and the vector pUC18 (including green fluorescent protein) were digested by the restriction enzymes Bamhâ… and Sacâ… . The long segment of pBI121 and the segment of green fluorescent protein were recycled, and then were connected together. The positive clone was got. It was proved that the gfp (green fluorescent protein) had been connected with the vector pBI121 by PCR and enzyme digestion.