Development of Renewable Biofuels Technology by Transcriptomic Analysis and Metabolic Engineering of Diatoms

0903712希尔德布兰德该奖项由2009年美国复苏和再投资法案(公共法律第111-5条)资助。该项目的目标是开发硅藻的代谢工程方法，通过可控地操纵细胞内过程而不是外部环境条件来诱导脂肪积累，并操纵细胞内碳在脂肪和碳水化合物合成之间的分配，以实现丰富的生物量和脂肪积累。有四个具体的目标：1)对硅、氮和硒限制引起的脂肪积累的硅藻进行比较转录分析，以确定参与控制脂肪积累和碳分配的关键调控步骤。2)对硅藻细胞进行代谢工程，使之能够通过简单的操作来抑制硅的运输，作为一种人为地使细胞饥饿以触发脂质积累的方法。3)对细胞进行代谢工程，使其能够通过简单的操作来抑制硝酸盐的运输，作为一种人为地使细胞饥饿以获得氮以触发脂质积累的手段。4)对细胞进行代谢工程，以改变碳分配，以实现丰富的脂肪积累和高生物量生产，而不需要营养限制。智力价值：该项目的意义在于，通过使用可操纵的细胞内过程而不是依赖于可变的环境条件，它将能够更好地控制用于生物燃料生产的硅藻中的脂肪积累，并且它可能使脂肪在正常生长条件下积累。在微藻中，脂肪积累通常发生在营养限制条件下，这种条件受环境变异性的影响，从而阻止了高生物量积累。将脂质诱导的控制转移到细胞内过程将消除这种可变性，识别控制细胞内碳分配的关键调控步骤与代谢工程相结合，应该能够在非限制性营养生长条件下将碳更多地分配到脂质中。这些方法预计将对可再生生物燃料技术的发展产生重大影响。更广泛的影响：本科生和高中生参与这个项目将使他们能够体验到“动手”的实验室研究。这对年轻人的思想发展至关重要，能够欣赏发现的实际和动态方面，而不是仅仅通过演讲或阅读吸收信息。这一点的重要性怎么强调都不为过，因为它使人们认识到任何成功的努力都必须应用于努力和承诺的程度。这些学生将作为当地培训项目的一部分参加，这些培训项目强调未被充分代表的少数民族的参与。拟议研究的社会效益将通过发展硅藻作为可再生燃料来源的技术应用来实现。潜在的好处包括减少二氧化碳的产生和随后对气候变化的好处，减少对石油进口的依赖，以及在这个新兴的技术行业创造新的就业机会。

0903712 HildebrandThis award is funded under the American Recovery and Reinvestment Act of 2009 (Public Law 111-5).The goal of this project is to develop metabolic engineering approaches for diatoms to enable induction of lipid accumulation by controllable manipulation of intracellular processes rather than from external environmental conditions, and to manipulate carbon partitioning within the cell between lipid and carbohydrate synthesis to enable both abundant biomass and lipid accumulation. There are four specific objectives: 1) To perform comparative transcriptomic analyses in diatoms of lipid accumulation resulting from silicon, nitrogen, and selenium limitation, to identify key regulatory steps involved in controlling lipid accumulation and carbon partitioning. 2) To metabolically engineer the diatom cell to enable repression of silicon transport by simple manipulations, as a means to artificially starve the cells for silicon to trigger lipid accumulation. 3) To metabolically engineer the cell to enable repression of nitrate transport by simple manipulations, as a means to artificially starve the cells for nitrogen to trigger lipid accumulation. 4) To metabolically engineer the cell to alter carbon partitioning to enable abundant lipid accumulation along with high biomass production, without the need for nutrient limitation. Intellectual Merit: The significance of this project is that it will enable greater control over lipid accumulation in diatoms for biofuels production by using manipulable intracellular processes rather than depending on variable environmental conditions, and it will possibly enable lipid accumulation under normal growth conditions. In microalgae, lipid accumulation generally occurs under nutrient limiting conditions, which are subject to environmental variability, and which prevents high biomass accumulation. Transferring control of lipid induction to intracellular processes will eliminate the variability, and identifying the key regulatory steps involved in controlling carbon partitioning in the cell coupled with metabolic engineering should enable greater partitioning of carbon into lipids during non-limiting nutrient growth conditions. These approaches are expected to have a substantial impact on the development of renewable biofuels technology. Broader Impact: The involvement of undergraduate and high school students in this project will enable them to experience "hands on" laboratory research. This is critical in the development of a young person's mind, to be able to appreciate the practical and dynamic aspects of discovery, as apposed to just assimilating information via lectures or reading. This importance of this cannot be overemphasized, because it develops an appreciation for the level of effort and commitment that must be applied to any successful endeavor. The students will participate as part of local training programs that emphasize the involvement of underrepresented minorities. The societal benefits of the proposed research will be realized through the technological application of developing diatoms as a source of renewable fuels. The potential benefits include reduced CO2 production and subsequent benefit to climate change, decreased dependence on oil imports, and establishment of new jobs in this emerging technology industry.