Controlling Heterogeneity in Plant Cell Culture and Secondary Product Accumulation through Metabolic Analysis and Modeling of Population Behavior

通过代谢分析和群体行为建模来控制植物细胞培养的异质性和次生产物积累拟议的研究项目集中在使用新方法来研究和表征植物细胞培养的异质性。具体目标是：(1)利用多参数流式细胞术获得红豆杉种群的基线信息，以表征代谢异质性，并建立种群平衡方程模型，将代谢数据与过程尺度信息关联起来；(2)利用新型荧光活化细胞分捡（FACS）技术，基于紫杉醇积累分离亚种群，并表征代谢异质性和亚种群中的代谢异质性；(3)基于紫杉醇积累对亚种群进行分类和恢复，分析生长和紫杉醇积累的长期稳定性，建立具有预测种群行为能力的种群平衡方程模型。智力优势：本研究的结果将提供红豆杉异质性的详细特征，然后可用于设计有效的策略来稳定和优化紫杉醇积累。这是非常有价值的，因为未来紫杉醇的供应很可能在很大程度上取决于植物细胞培养技术的使用。更广泛的影响：这项研究的结果有可能在创造新的方法（例如，单细胞分析和单个植物细胞的分选）和研究红豆杉和其他植物系统的研究人员的方法方面对科学界产生重大影响。所有参与该项目的人员都将接受实验和建模方法的培训，并将积极参与一项新的指导计划。代表性不足的学生将通过国家科学基金会赞助的教育计划被招募。

Controlling Heterogeneity in Plant Cell Culture and Secondary Product Accumulation through Metabolic Analysis and Modeling of Population BehaviorSusan RobertsUniversity of MassachusettsCBET-0730779The proposed research project centers on the use of novel approaches to study and characterize plant cell culture heterogeneity. The specific aims are to: (1) obtain baseline Taxus population information using multiparameter flow cytometry to characterize metabolic heterogeneity and develop population balance equation models to correlate metabolic data with process scale information, (2) isolate subpopulations based on paclitaxel accumulation using novel Fluorescence Activated Cell Sorting (FACS) techniques, and characterize metabolic heterogeneity and in the subpopulations, and (3) sort and recover subpopulations based on paclitaxel accumulation, analyze long-term stability in terms of growth and paclitaxel accumulation, and develop population balance equation models with the ability to predict population behavior. Intellectual Merit: The results from the proposed research will provide a detailed characterization of Taxus heterogeneity, which can then be used to design effective strategies to stabilize and optimize paclitaxel accumulation. This is of great value because future paclitaxel supply is likely to depend significantly on the use of plant cell culture technology. Broader Impacts: The results of this research have the potential to significantly impact the scientific community in the creation of new methodologies (e.g., single cell analysis and sorting of single plant cells) and approaches for investigators studying Taxus and other plant systems. All personnel participating in this project will be trained in both experimental and modeling approaches and will actively participate in a novel mentoring program. Underrepresented students will be recruited through the NSF-sponsored educational program.