| Tetrastigma heleyanum Diels et Gil(T.heleyanum) is perennial plant of vines Vitaceae, and is rare and endemic herbs in China. At present, the research of T.heleyanum is more in pharmacognosy, chemical constituents, pharmacological effect and clinical application. However, few studies have been done on tissue culture. A comparatively perfect system of tissue culture and rapid propagation had been established in T.heleyanum. This system could be used for field production further.The optimized system were as follows:In contamination control, shoot induction medium containing0.3g/1fungistat and1mg/l ampicillin could effectively curb the explants contamination and the contamination percentage decreased10%-50%; The concentration and combination of different plant growth regulator had a significant effect on shoot induction and multiplication. The results showed combination of0.8mg/l6-BA and0.05mg/l NAA was suitable for shoot induction and the shoot induction percentage reached100%. But for shoot multiplication, the combination of2.0mg/l6-BA and0.1mg/l NAA was the optimum and the multiplication coefficient reached8.1/2MS and WPM were the suitable shoot induction basal medium, however, B5and WPM basal medium were suitable for shoot multiplication. The combination of1mg/l NAA and1mg/I IBA was benefit for rooting. Not only the rooting percentage reached100%but also the roots were stronger and the number of roots was more. The whole process of tissue cuture in T.heleyanum was about90days from explant sterilization to plantlet. Obvious swell tuberous could be obseverd after T.heleyanum transplanted for four months.Piriformospora indica is a fungus that can widely colonized in a variety of plants. The colonization of P. indica confers plants strong growth promotion, induces disease resistance to abiotic and microbial pathogens, promotes nutrition absorption and increases yield and quality. The agricultural potential of P. indica is deduced from the wide range of mutualistic symbioses plants and obvious stimulation effects. In this study, P. indica began to colonize in T.heleyanum roots at the sixth day after inoculated, which was proved by the results of PCR and trypan blue staining. P.indica colonization promoted T.heleyanum growth and development. And the stem length, dry weight and fresh weight both were obvious increased compared with controls. The auxin leve in the T.heleyanum buds was higher than uninoculated controls during10days after inoculated, but the auxin leve of roots was opposite results. Furthermore, the gene expression of vacuolar-type H+-ATPase subunit B2, which is releated to cell wall acidication by auxin leve regulation, was down-regulation in T.heleyanum root. While, the polar transport related genes PIN3expression was up-regulation. |
