Study On Technique System Of Tissue Culture And Characteristics Of Cultures From Style In Crocus Sativus L.

Crocus sativus L, known as saffron, is a perennial herb of Iridaceae family with high medical value and also a kind of precious raw material of chemical dyestuffs and flavor. But the exploitation of saffron is restricted by its low yield with traditional culture technics and production methods, causing high cost for the lack of resource.Therefor, the researches of prodcuting efficiently medical tissue and active compounds using the technology of tissue culture have already been paid attention to.As only its dried stigmas can be used for medicine and spice, the stigma,style and stamen were used in primalry culture ,while the cultures from style in subculture to study the changes occurred to the proliferation,organ differentiation and characteristics in the process of primary culture and subculture, and other affecting factors. A technique system were primarily established through discussing the optimize conditions of flowering, desiccation process for the culture product and chemical element analysis. The results are as follows:1. This paper investigates the varied effects of explants, different treatments and inoculations to the results in primary culture. It indicated that the best result with less pollution and higher inducing rate can be obtained with the adoption of the improved one-step Sterilization method, suitable cut fashion and inoculation. And the styles at each development stages were best suited for the regeneration of callus among three explants.2. Auxin was an important factor affecting the induction of callus in primary culture. And lower concentrate NAA is favorable for the formation of callus within a certain rang of concentration(within2.0-8.0 mg/L).Though 6-BA effects less than NAA, the interaction between them were significant. 2.0mg/L of NAA associated with 1.0mg/L of BA would be most beneficial for the induction of callus. Furthermore, hormone conditions have significant effect on the exterior characteristics, such as colors, homogeneity and the texture of callus. Good exterior conditions and a fast-growing could be achieved by the following conditions: MS+6-BA 5.0 mg/L+ NAA 2.0 mg/L + 2,4-D 2.0 mg/L, thus primarily establishing the technique system .3. Floral differentiation and flowering in vitro from callus were also observed in the primary culture. The stamen and style at the same stage had different ways of flowering under the media and cultural conditions. The styles firstly differentiated to be callus which would then differentiated into flower or other organs, while the stamens directly differentiated into flower or its organs. The stigma can't form any floral organs. The relatively mature style (A2) formed the comparative complete flower with stigmas (or stigma-like structures) + petal-like structures and ease to flowering in vitro while the younger style (A1) only regenerated the petals or petal-like structures. Neither of those two explants in the different development stages would differentiate into stamen. The ratio of BA/NAA plays a leading role in differentiation types of floral organs, that is, a high ratio of 6-BA/NAA is helpful for callus to differentiate the stigma-like structures, NAA is the optimalizing auxin in the culture of style which followed by IBA and 2,4-D .4. Many types of morphological differentiation were observed in subculture of the style.The study shows that the accumulation of chemical composition had been related to morphological characters to a certain degree while hormone had played a significant role. The condition for principal component production had been conclusively figured out . what's more, the proliferation, increment and exterior conditions presented unstability as the passage number increased, with the proliferation changes from declining to improving and the exterior conditions with no regularity except being placed in the media condition (MS+6-BA5.0mg/L+ NAA5.0mg/L) when the organ differentiation was relatively stable.5. Pre-treatment method determined in the study was beneficial to improve the quality of exterior conditions of the cultures. The temperature adopted in the pre- treatment remarkably effected the kinds and contents of chemical compositions.It was also showed that the different types of cultures and extracting methods would result in different chemical components with different characteristic peaks. Compounds contained in the cultures of style are saccharides, flavones, alkaloids, saponins, phenolic compounds and tannins, organic acids, anthraquinones, coumarin and lactones, steroids or diterpenes,etc. And the amount of soluble sugar and total flavonoids in these cultures were the same as they were in natural stigma. Among those chemical components of more 17 kinds, two of them at least could be found as in a natural stigma.