Studies On Cell Differentiation And Physiologic Mechanism Of Scale Culture Cardiocrinum Giganteum

This study dealt with the scale of Cardiocrinum giganteum of as materials. And treating. the callus induction as the basic experimental system. By adding different types of exogenous hormones, observing the Cardiocrinum giganteum's callus induction, the process of differentiation of tissue morphology changes. Analyzing the amount of endogenous hormones, nucleic acid, soluble protein content of the callus tissue, which had been treated with different hormones, discovering the rule of changes of aperoxidase (POD), phenylalanine ammonia-lyase (PAL), protease's activity. And also doing study on the peroxidase and esterase isozymes. Learning the mutual relations amoung, the cell differentiation, organogenesis, the physiologically biochemical basis, and the culture conditions, in the process of Cardiocrinum giganteum's tissue culture. The main conclusions as follows:Firstly, the studies on the physiological and biochemical conditions in the callus induction phase indicated that the MS medium containingNAA0.1mg/L plus 6-BA1.5mg/L can induce the synthesis of endogenous hormone and cytokinin (iPAs. ZRs). IAA, iPAs influence the cell growth start and the formation of callus very much. In the callus induction process, there were tight relation amoung the DNA, RNA and protein. POD, PAL, protease activity showed some changes, and the isozyme emerged some abnormal bands.Sceondly, different types and different concentrations of hormone combinations made differences on the growth effect, organ occurrence of Cardiocrinurn giganteum's callus. The callus can grow well in the MS medium containing NAA0.1mg/L plus 6-BA1.5mg/L with the various combinations of estrogen treatments. The callus grew well in both of the MS medium containing NAA0.1～3.0mg/L plus 6-BA1.5mg/L and the medium containing NAA0.1mg/L plus 6-BA0.0～4.0mg/L, also had adventitious buds. High concentrations of NAA or 6-BA inhibited the adventitious buds. The callus grew best in the MS medium containing NAA0.1mg/L plus KT0.2～1.5mg/L, and the adventitious buds and the roots had been induced much. The callus grew slowly in the medium containing NAA0.1mg/L plus 6-BA1.0～2.0mg/L and KT0.5～1.0mg/L, but there was no organogenesis. Considering the callus growth and cell division, the MS medium containing NAA0.1mg/L plus KT0.2～1.0 mg/L was the best.Thirdly, according to the study on the relationship between different types and different concentrations of hormones and appearance changes, the conclusions are as follows: The adventitious buds were related with the level of the IAA. There was no adventitious bud, if the level of IAA was low. And there all had adventitious buds with addition of 6-BA and KT, when IAA content reached a certain level above. Meanwhile the producing of adventitious buds needed a period of reducing the amount of IAA. But in different types of hormone treatment, the number of adventitious buds was not equal to the speed of reducing of IAA. The producing of adventitious buds in all the treatment was accompanied with the content reducing of CTK. The root division was affected by the IAA and iPAs, and the adventitious root formation needed the low level of IAA and CTK.Fourthly, the study on the DNA content in callus of different types and concentrations of estrogen treatment can show that nuclide was a standard of changes of Cardiocrinum giganteum's cell. RNA increased, while DNA content dropped, before the callus tissue differentiation. The increase of RNA content in the callus tissue marked the material synthesis of cell differentiation. In the process of bud primordium formation, RNA decline. And that was the sign of cell differentiation. The start-up of root primordium formation needed metabolism of DNA reaching a high level.Fifthly, the study on the activity of soluble protein content, protease, Peroxidase (POD), phenylalanine ammonia-lyase (PAL) during the process of callus differentiation showed those as follows:The amount of protein, and protease activity were related to the callus growth, but not as the sign of cell differentiation. The activity of POD and PAL was tight to the formation of organs, were varied in different treatments. The decrease of POD's activity of callus in different treatments( NAA and KT), affected the producing of adventitious buds. KT was key factor to root differentiation, and the adventitious buds differentiation completed after the second activity peak of 6-BA. In all the treatments, PAL activity was decreasing while adventitious buds forming. In the treatment(KT), the second activity peak was consistent with the root primordium formation.At last, the differences of POD isozyme reflected on the enzyme bands. There was no special band appeared in the occurrence of the adventitious buds and roots. EST isozyme two focus areas. NAA can enhance the second enzyme band, inhibited the enzyme activity of the first enzyme band, and made the final enzyme band disappeared. 6-BA can greatly enhance the enzyme activity of two concentrated areas, but KT made the enzyme activity of those two bands weakened. The callus organogenesis of Cardiocrinum giganteum was related to the mutual effects of all the enzyme bands.