雨生红球藻信号物质的研究

IOCAS-IR > 海洋环流与波动重点实验室

	雨生红球藻信号物质的研究
	孙艳妮
学位类型	博士
	2001
学位授予单位	中国科学院海洋研究所
学位授予地点	中国科学院海洋研究所
学位专业	海洋生物
关键词	红球藻旧液抑制物质提取物游动细胞不动细胞
摘要	虾青素作为饲料添加剂、天然食用色素和生物保健品活性成分，具有很好的应用前景，目前国际市场上价格昂贵，供不应求。雨生红球藻是虾青素的良好资源。但是，由于种种原因限制，红球藻细胞培养仍然不很理想，成为限制工程规模放大和开发该生物资源的瓶颈。那么，是否在光照、温度、pH、营养盐等传统影响因子之处，还有调节红球藻细胞生长的重要因子？目前国内外尚不清楚。在国家自然科学基金和国际科学基金资助下，本论文围绕红球藻是否产生调节细胞周期信号物质的语题，利用红球藻处理过的旧液重新进行该藻培养，去排除其它影响因子的基础上，初步结果表明红球藻确实产生某种物质，抑制自身生长。主要研究结果总结如下：1、在静置培养时，通常难于直接观察到旧液对藻细胞的抑制现象，这主要是由于其它因素（如无机碳等）的存在掩盖了抑制物的作用。因此，不难理解为什么以往研究大多忽视抑制物问题。2、在通气和加富二氧化碳通气培养时，旧液对藻细胞的抑制现象就比较容易发现。该物质可溶于二甲基亚枫等有机溶剂的物质、可以用乙酸乙酯从旧液中提取出来。尽管旧液中存在碳酸根和碳酸氢根等无机碳，但它们不是抑制性信号物质。3、该信号物质的产生与藻培养的时间也有一定关系，藻培养的初期并不大量积累，但在培养过程（特别是在培养的中、后期）逐渐形成，并积累在旧液中。这就可以从一个侧面阐述为什么常常难以长时间维持红球藻细胞的连续培养。4、该信号物质的产生与处理藻细胞密度有关，细胞密度越大，红球藻产生的抑制信号物质就越多。因此，高密度培养和超高密度培养中，抑制物是较为严重地限制红球藻细胞生长的问题之一，可能也是微藻（甚至细胞）培养中值得研究和关注的话题。5、红球藻游动细胞是抑制性物质产生的主要阶段，随着游动细胞转化成不动细胞，该信号物质并没有明显增加的迹象。6、尽管目前还不清楚抑制如何调节红球藻细胞周期，但是，在旧液中或有抑制性物质存在条件下，红球藻游动细胞分裂受阻，数量下降，同时不动细胞数量增加。推测抑制物有抑制游动细胞分裂，和促进游动细胞向不动细胞转化的作用。同时，有关DNA含量和DNA倍性结果也初步表明，红球藻游动细胞DNA复制过程没有受影响，但随后的细胞原生质分裂过程受阻。至于红球藻类细胞的原生质分裂受阻，和该藻细胞周期进入不动细胞阶段之间可能存在的机制，还有待进一步探讨。总之，本论文证明了红球藻在生长过程中向胞外释放降低细胞增长和诱导细胞转化的信号物质，这是细胞生长的又一个限制因子，为提高细胞培养密度和深入了解细胞周期调控机制提供了基础数据。
其他摘要	Astaxanthin, as a good colorant additive and a bioactive substance, has great commercial potential. The limited output of astaxanthin nowadays can't meet the increasing demand worldwide. Therefore, much attention has been given to a unicellular green alga, Haematococcus pluvialis, in recent years due to its ability to accumulate large amount of astaxanthin. The possible effects of different traditional factors, like light (light regime and light intensity), temperature, nutrients (N, P, C, Fe, trace mental elements), vitamins, salinity and pH on the cell growth of this alga have been studied in number of laboratories and countries with a lot fruitful results. Unfortunately, the cell growth rate is still poor, the cell density of the culture is relatively low and the cell growth rate is still poor, the cell density of the culture is relatively low and the cell cycle is not well regulated. We suspected a new factor (auto-signals) maybe impress the cell growth of the motile cells, induce the cell transformation from the motile cells to the non-motile cells, and limit the scale up of culture this green alga. However, little is known about this interesting subject both in theory and in practice. Our aims of this thesis focus on above issue. Some positive progresses have been achieved thank to the financial supports from IFS (International Foundation for Sciences) and NSFC (National Nature Sciences foundation of China). The main results obtained are listed as below: 1. Auto-signals did exist in the old culture supernatant and pay a significant inhibitory role in regulating cell cycle. The inhibitory effect of old culture supernatant would be easily detected especially in aerated culture and CO_2 enriched aerated culture, but not in static culture. In the latter culture mode, other factors, such as inorganic carbon, cover up the inhibitory effect, which maybe the reason why this important inhibitory phenomenon has long been neglected in many early works. 2. The auto-signals in the old culture supernatant would be extracted by ethyl acetate and dissolved in dimethyl sulphoxide. HCO_3~-, CO_3~(2-) and other inorganic carbon were not connected with auto-signals although they really existed in the old culture supernatant and affected the cell growth too. 3. Auto-signals were produced from Haematococcus itself and excreted into the culture medium because the production of auto-signals was culture-time dependent. No inhibitory effect effect was detected at the early culture stage. The longer the culture supernatant was treated by the alga, the more the auto-signals accumulated in the culture supernatant, and the heavier the cell growth was inhibited. This maybe the right reason why H. pluvialis would not be easily cultivated for long. 4. The accumulation of auto-signals was connects with cell density too. More signals were produced in the high cell density culture. Therefore, auto-signals became one of the factors serious blocking cell growth in high even ultra-high culture of H. pluvialis, and need pay much attention to study deeply. 5. Auto-signals were mainly produced by the motile cells of H. pluvialis. When the motile cells transformed into the non-motile cells, no evident showed a further increase of auto-signals in the old culture supernatant. 6. Although the mechanism of the auto-signals regulating the cell cycle has not clear yet, we still concluded a decrease in the motile cell parallel with a increase in the non-motile cell when H. pluvialis exposed to old culture supernatant or its crude extracts. Therefore, we suggested that auto-signals block the motile cell division and accelerate cell transformation from the motile cells to the non-motile cells. The DNA data of the motile cell showed that the process of DNA replication in the motile cells was not affected by auto-signals, but the process of the plasm division was heavily inhibited. More DNA data of the non-motile cell are required to better understand the story of cell cycle regulation in this alga caused by auto-signals. In brief, H. pluvialis released auto-signals into the culture medium, which as a new limiting factor feedback inhibited its cell growth and induced cell transformation.
页数	39
语种	中文
文献类型	学位论文
条目标识符	http://ir.qdio.ac.cn/handle/337002/689
专题	海洋环流与波动重点实验室
推荐引用方式 GB/T 7714	孙艳妮. 雨生红球藻信号物质的研究[D]. 中国科学院海洋研究所. 中国科学院海洋研究所,2001.

条目包含的文件
文件名称/大小	文献类型	版本类型	开放类型	使用许可
LW005529.pdf（2830KB）			限制开放	--	浏览