Establishment Of Agrobacterium Tumefaciens-mediated Transformation Of Somatic Embryos System And Studies On Atft Gene In Hevea Brasiliensis

Hevea brasiliensis Muell.Arg is a perennially cross-pollinat and high heterozygous woody plants. Using conventional breeding methods, a new breed nurturing need20to30years. With the rapid development of biotechnology, genetic engineering, somatic mutation and somatic cell hybridization technology continue to mature and have highly efficient directional advantages, which are an effective way to shorten the breeding cycle and creats favorable conditions for improved varieties of rubber tree.The secondary somatic embryogenesis as receptor material to establish the genetic transformation of rubber tree. Somatic embryos of RY7-33-97as recipient, study acetosyringone, infection time, co-culture days and antioxidants have effect on somatic embryos. Establishment of agrobacterium tumefaciens-mediated transformation of somatic embryos system. On this basis, studies Arabidopsis FT transgenic by screening kanamycin resistance, secondary embryogenesis and GUS histochemical staining positive embryoid. The main results as follows:1. According to GUS transient expression rate, use single-factor experimental design five factors as acetosyringone, infection time (static and vacuum), co-culture days and the antioxidant(silver nitrate, sodium thiosulfate, halfcystine and dithiothreitol); orthogonal experimental design four antioxidants with silver nitrate, sodium thiosulfate, halfcystine and dithiothreitol, and orthogonal experimental design acetosyringone, vacuum infection time and co-culture days, study on Agrobacterium-mediated transformation somatic embryos genetic in Hevea brasiliensis. Establishment of Agrobacterium tumefaciens-mediated Transformation of Somatic embryos system conditions as follows: Agrobacterium concentration OD600=0.3～0.4, infection8min with vacuum, co-cultured with anti-oxidants (Na2S2O3,1.5mmol/L+AgNO315mg/L+DTT1.5mmol/L), co-cultured five days.2. According to GUS Stable expression rate, the single-factor experimental design acetosyringone, still infection, antioxidants and co-culture day on AtFT gene genetic transformation rubber tree. In this way, the total number of230embryos, transformed embryo51, a total of291secondary somatic,59GUS dyeing positive, of which14dark blue and37light blue.