In Vitro Culture And Analysis Of Changes Of Endogenous Hormones In Wuyimingcong Cultivar Group Of Camellia Sinensis L.

In this experiment the adult stem nodes were used as materials to carry out in vitro culture of Wuyimingcong cultivar group such as the early-sprouting line Jin Fenghuang, the middle-sprouting line Jin Suoshi and the late-sprouting cultivar Rou Gui. The aseptic lines were first established, and then the studies on the bud proliferation and rooting were conducted, and finally the difference among the three cultivars was compared. In the meanwhile, the basic condition parameters of extraction and determination on endogenous hormones in tea trees were investigated, and the changes of the endogenous hormones in both explants and in vitro materials were analysed by HPLC. Morover, the genetic stability of the materials of pre and post in vitro culture were performed by RAPD analysis. The main results were as follows:1. In vitro culture of adult stems of Wuyimingcong cultivar group. Experiment with different sterilization methods, as well as cutting mode and different light intensity, the different types of growth hormone, GA₃ concentration of the different hormone levels, as well as salt and sugar concentration on Wuyimingcong axillary buds were induced, and also compared the different hormone levels and the proliferation of different types of salt and and the impact on the root. The results showed that the most suitable explants were two-leaf-located stem materials in spring and one-bud-two-leaf period, the most appropriate disinfection method was using 0.1 % mercuric chloride disinfectant for 9～11 minutes which depended on different cultivars as well as their tender extent, and medium MS+2.0 mg·L^-1BA+0.5 mg·L^-1IAA+0.5 mg·L^-1 GA₃ was suitable for initiation culture of variety Rou Gui, medium 1/2MS+1.0～1.5 mg·L^-1BA+0.2 0.8 mg·L^-1IAA+0.2 mg·L^-1 GA₃ reached fair propagation effects on subculture of Wuyimingcong, and the ordinal adaptability to medium was Jin Fenghuang, Jin Suoshi and Rou Gui, medium 1/4 MS+4 mg·L^-1 NAA containing lower minery and higher auxin concentrations was beneficial to rooting. The average propagation coefficient was 3.00 by propagation culture, and under this culture system it might be predicated that 729 axillary buds be regenerated and propagated from one bud within a whole year, which opened up broad prospect of applications to tea production.2. Changes of endogenous hormones in Wuyimingcong by HPLC.(1)On investigating the endogenous hormones in tea trees, the selected wavelength of GA₃ was 207 nm, and that of IAA, ABA and ZT was respectively 280 nm, 280 nm and 254 nm, the mobile phase for the analysis of GA₃ was methanol- acetonitrile-KH₂PO₃ buffer system (15:15:70, v/v, pH 3.5), the mobile phase for the analysis of both IAA and ABA was methanol-acetonitrile-KH₂PO₃ buffer system (20:20:60, v/v, pH 3.5) , the mobile phase for the analysis of zeatin methanol - acetonitrile -phosphate buffer (15:15:70, v/v, pH 7.0). (2) On investigating endogenous hormones in both explants and plantlets, the between-changers contents and content variation ranges might be relevant to genetic characters of the cultivars, and GA₃ was one of the indispensible endogenous hormones during the growth and development of tea trees. The two-leaf-located leaves in different growth phases, different leaf-located leaves in the same growth phases and two-leaf-located leaves grew vigorously, whose variation ranges of endogenous hormones were relevatively larger, moreover, spring was the suitable season for the growth of bud seedlings. The propagation effect of medium adding exogenous hormones BA, IAA and GA₃ was better than free hormones, and under that circumstances the content of IAA and ZT increased while the content of GA₃ declined gradually, where the content of ABA varied little.3. Genetic stability test on in vitro plantlets of Wuyimingcong by RAPD analysis. The main results showed that it varied between pre and post in vitro culture of Rou Gui by 4.13 %, From the production point of view, the existence of 4.13 percent in the fields of plant variation did not affect the quality of tea products, which was stable so that meristems were reasonably used for in vitro culture. On the other hand, the purity of in vitro plantlets was higher while the DNA concentrations of plantlets and explants were almost equivalent and of stable contents, both convenient for material access and DNA extraction.