Method for stably culturing taxus chinensis plant cell seeds

Abstract

The invention discloses a method for stably culturing taxus chinensis plant cell seeds, and aims to provide a method which is capable of being used for stably culturing the taxus chinensis plant cell seeds; and for the method, the advantages of being economically feasible, simple and controllable in process, and low in cost are realized. The method comprises the steps of: (1) sampling and detecting subculturing seeds; (2) adjusting a final biomass of the seeds to 5-10g / L; (3) increasing the initial sugar degree of a culture solution to 15-25g / L; (4) increasing the initial electric conductivity of the culture solution to 2.5-3.5ms / cm; (5) controlling an initial biomass of the culture solution to be in the range of 2.5-3.5g / L and the initial pH (potential of Hydrogen) to be in the range of 5.0-6.0, stirring and carrying out seed subculturing; and (6) when the subcultured seeds in the step (5) meet the requirements that A, the final sugar degree of the culture solution is greater than 8g / L or B, the final electric conductivity of the culture solution is greater than 2.0ms / cm or C, the final pH of the culture solution is less than 6.0, starting next seed subculturing. The method belongs to the technical field of seed culturing.

Description

technical field [0001] The patent of the present invention relates to a method for stably cultivating cell seeds, in particular to a method for stably cultivating yew plant cell seeds, which belongs to the technical field of seed cultivation. Background technique [0002] Paclitaxel (Taxol) is a diterpene alkaloid isolated and extracted from the bark of the yew plant for the first time. expensive. [0003] At present, there are three main sources of paclitaxel raw materials: one is to obtain paclitaxel from the branches and leaves of Taxus chinensis, but due to the slow growth of Taxus chinensis, the production cycle is as long as 3 to 5 years, and the content of paclitaxel in the branches and leaves is low, resulting in the extraction of paclitaxel. high cost. The second is to semi-synthesize paclitaxel through precursors such as Baccatin Ⅲ and 10-DAB, but the semi-synthesized precursors also come from planting branches and leaves, which are also limited by the source of ...

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Problems solved by technology

[0003] At present, there are three main sources of paclitaxel raw materials: one is to obtain paclitaxel from the branches and leaves of Taxus chinensis, but due to the slow growth of Taxus chinensis, the production cycle is as long as 3 to 5 years, and the content of paclitaxel in the branches and leaves is low, resulting in the extraction of paclitaxel. high cost

The second is to semi-synthesize paclitaxel through precursors such as Baccatin Ⅲ and 10~DAB, but the semi-synthetic precursors required also come from planting branches and leaves, which are also limited by the source of raw materials for extraction, and still cannot meet the large demand for paclitaxel in clinical and experimental studies

[0004] China, the United States, Canada, South Korea, Japan and other countries have conducted a lot of research on the production of paclitaxel using yew plant cell culture technology, but the research hotspots of scientists are mainly concentrated on the high yield of paclitaxel. So far, no set of yew cell seeds has been proposed for long-term stable succession. The technical scheme of subculture, due to the great difference in seed quality and growth state, the growth is slow, red or brown substances will be continuously produced during the growth process, and the technology of subculture often occurs due to allergies, redness or senescence, death and browning, and fails Difficulties, resulting in poor stability of paclitaxel content between batches when plant cells are cultured to produce paclitaxel, which is a common technical problem faced by the large-scale cultivation of yew plant cells in the industrial production of paclitaxel. The technical research on the production of paclitaxel, so far, few research teams have entered the pilot test and industrialization stage, and there is no way to cultivate yew cell seeds with stable quality and growth state, which is one of the main reasons for the failure of cell culture

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