Study On Antisense TCS Gene By Agrobacterium-mediated Transformation To Mature Embryo Of Tea Plant

Tea (Camellia sinensis (L) O. kuntze) is primary target for the application of genetic engineering to improving agronomic characteristic. In this paper, some of the excellent regeneration system in tea plant were established . The mature embryos used as receptor to be transformed by Agrobacterium tumefaciens with antisense TCS gene and the parameter of transformation were optimized by GUS transient expression. The mature embryos were cultured on selection media with Kan. The main results had been obtained as follows:1,Cultivation of mature embryosBy establishment of regeneration systems, mature embryos induced young plant firstly, which is the base of heredity transformation on tea plant. In the cultivation, tea plant genotype, concentration of hormone in the medium are the key factors that influence of callus and plant regeneration. The optimize concentration of 2,4-D and KT is defferent, as for as Long jing 43# and Nong kang zao, optimal medium was MS supplemented with 2 mg/L 2,4-D and 0.5mg/L KT, but as for Chu ye zhong, optimal medium was MS supplemented with 2mg/L 2,4-D and 1.0mg/L KT. Genotype mainly influences the regeneration frequency of callus formation, 14.2% buds regeneration frequency can form by Long jing 43#, 8.5% buds regeneration frequency can form by Nong kang zao, 3.3% buds regeneration frequency can form by Chu ye hang , and there is 40% root formation frequency. By induced callus track, mature embryos needed more one year cultivate integrate tea plant. The MS medium with 2 mg/L 6-BA was suitable for mature embryo to culture young plant directly. Root formation frequency is 86%. Mature embryos cultivated stuggy tea plant directly which need more half one year.2,Establishment of Agrobacterium-mediated transformation systemsThe conditions of infection were optimized by GUS transient expression. The results indicated that concentration of Agrobacterium is 0.6 , the efficient density of AS is 500μmol/L and the time of co-cultivation is 5 days.3,Selection and ResultSelection system of high efficiency was determined by different concentration of Kan. Mature embryos growth and rooting concentration were 50 mg/L and 75 mg/L respectively. Effective concentration of Carb which restrains Agrobacterium tumefaciens is 400mg/L. Under the selection of Kan, there are 10 young tea plant, which were checked by RT-PCR technology.