稀土胁迫斜生栅藻油脂积累机制研究

The exhaustion of fossil fuels since its non-renewal, the problems of environmental pollution and greenhouse effect are promoting the development of renewable and environmentally friendly energy sources all over the world. Biodiesel could be used directly as a substitute for fossil energy, but the serious shortage of raw materials has been the main factor of limiting the development of its mass production. In recent years, oil-rich microalgae have been suggested as a very good candidate used for biodiesel production as the raw material source. But the high cost is the bottleneck of its industrialized production. Except the microalgae strain selection and cultivation process improvement, themicroalgae oil content elevation is also a key technology for saving production cost..In this project,Scenedesmus obliquus strains were selected with higher growth rate and higher lipid production. Separation, screen, identification and optimization conditions for growth were studied. At the same time, wastewater in tailings reservoir from BaoGang was used to culture oil-rich microalgae, and the effects of Cerium on growth of oil-rich microalgae were investigated from physiology and biochemistry compositions, photosynthetic characteristics, and gene expressioncorrelation with lipid metabolism. Finally, the relationship between cell growth, lipid accumulation and the rare earth element existed in the environment were illuminated.There are very important theoretical promotion and practical guiding significance to improve the oil content of microalgae by genetic engineering and systems biologist techniques.

化石能源储量有限、不可再生，正在走向枯竭，且会带来空气污染。寻找可再生和环境友好的新能源，已成为各国的战略选择。生物柴油作为化石能源的替代品可直接使用，但制约其产业化的关键问题是原料不足。近年来，普遍认为富油微藻是未来制备生物柴油的最佳原料。但目前其工业化生产的瓶颈仍是成本高，除了藻种选择、培养工艺改进外，提高藻细胞油脂含量是最关键技术问题。.本项目选用本地筛选的生长快、含油高的斜生栅藻(Scenedesmus obliquus)，对其进行分子鉴定以及培养基营养盐和培养条件优化，利用包钢尾矿库废水进行培养驯化，分别从生理生化、光合反应以及油脂代谢相关基因表达差异分析等角度出发，研究稀土元素铈对藻细胞的胁迫过程，系统阐明能源微藻细胞生长、油脂积累与环境中稀土元素互作的响应机制，为通过基因工程和系统生物学调控手段提高能源微藻油脂产率提供理论及实验依据。

随着人类社会的发展，化石能源不断枯竭，微藻生物质能源备受关注，本项目以铈元素处理拟微绿球藻，测定铈元素对拟微绿球藻生物量、油脂积累，抗氧化系统以及光系统Ⅱ和油脂代谢相关基因表达的影响。首先对尼罗红染色法检测细胞内油脂含量的染色条件优化：二甲基亚砜浓度、预处理温度、尼罗红染液浓度，染色时间等进行单因素、正交实验，确定最佳染色条件；用铈元素处理拟微绿球藻细胞，低浓度铈元素对拟微绿球藻的生长并无影响，而高浓度的铈元素则抑制其生长。当铈元素浓度为52.6μmol•L-1时，稳定期拟微绿球藻油脂含量最大。通过透射电镜对细胞内铈元素进行定位，发现铈元素主要富集于液泡、内质网、细胞间隙等部位；在培养基中添加铈元素后，CAT、SOD两种酶的活性以及胞内MDA含量均有所增加，随着铈元素浓度增加，指数期细胞中叶绿素a含量先增加后降低，而在稳定期则不断降低。对于光系统Ⅱ供体侧吸收的能量增加，热耗散增加，受体侧接受的能量降低；将铈元素处理组以及对照组进行RNA-seq分析，发现铈元素胁迫下，拟微绿球藻的光合代谢途径受到很大的影响，这与差异表达基因的GO富集分析结果及前期铈元素胁迫导致光合效率降低的结果相一致。同时发现拟微绿球藻可能通过下调脂肪酸降解途径的相关基因的表达从而使油脂积累；丙酮酸脱氢酶复合体1、酰基辅酶A氧化酶、甘油激酶是差异基因中与油脂代谢相关的关键基因，经real-timePCR检测验证与RNA-seq结果相一致。本项目初步确定了铈元素对拟微绿球藻油脂合成的影响机制，从而为以拟微绿球藻作为生物柴油的原料提供理论支持。