Testable in silico Hypotheses for E.coli Growth

生物的基因型和表型之间的关系是生物学的基础。基于注释的基因组序列和生化信息，可以重建基因组尺度的代谢图谱，并且已经开发了计算机方法来解释和预测在给定生长条件下代谢图谱的最佳利用。利用已定义的大肠杆菌MG1655代谢基因型，实验上可测试的假设已经在硅片上制定，描述了初级碳源摄取速率、氧气摄取速率和细胞生长之间的定量关系。对大肠杆菌进行了实验，发现实验数据与在所考虑的实验条件下（醋酸盐、琥珀酸盐和苹果酸盐上生长），代谢网络支持大肠杆菌生长的最佳利用是一致的。这些重要的结果构成了拟议计划的基础。它由三个部分组成，1)继续为更广泛的碳源生成实验可测试的假设，并描绘这些生长条件下的基因表达模式；2)生成40个敲除菌株，并进行生长实验和基因表达研究，以表征其代谢行为；3)对表现出次优生长表型的大肠杆菌野生型和敲除菌株进行进化实验。此外，将对这些进化菌株在进化过程中和进化后进行基因表达研究，以确定进化过程中引发的分子事件。因此，实验程序的具体目的是确定一系列生长条件下的最佳相平面行为，并确定代谢基因表达模式是否与用于野生型和敲除菌株的预测途径一致。如果该计划成功实施和执行，我们将完善和提高我们对在所检查条件下大肠杆菌代谢基因型-表型关系的理解，这是一项具有明确基础意义的成就。

The relation between the genotype and the phenotype of an organism is fundamental to biology. Based on the annotated genome sequence and biochemical information it is possible to reconstruct genome-scale metabolic maps, and in silico methods have been developed to interpret and predict the optimal utilization of the metabolic map under a given growth condition. Using the defined Escherichia coli MG1655 metabolic genotype, experimentally testable hypotheses have been formulated in silico describing the quantitative relation between the uptake rate of a primary carbon source, oxygen uptake rate, and cellular growth. Experiments with E. coli have been performed and experimental data were found to be consistent with the optimal use of the metabolic network to support growth of E. coli under the experimental conditions considered, which were growth on acetate, succinate, and malate. These important results form the basis for the proposed program. It is comprised of three parts, 1) to continue the generation of experimentally testable hypotheses for a broader ranger of carbon sources and to profile the gene expression patterns for these growth conditions, 2) to generate 40 knockout strains and perform both growth experiments and gene expression studies to characterize their metabolic behavior and 3) to perform evolution experiments for wild-type and knockout strains of E. coli that exhibit sub-optimal growth phenotypes. Additionally, gene expression studies will be performed for these evolving strains during and after evolution to identify the molecular events that are elicited during the evolutionary process. Hence the experimental program has the specific aims of determining optimal phase plane behavior for a range of growth conditions, and to determine if metabolic gene expression patterns are consistent with predicted pathway used for both wild-type and knockout strains. If the program is successfully implemented and executed we will have refined and improved our understanding of the metabolic genotype-phenotype relationship for E. coli under the conditions examined, an accomplishment of clear fundamental significance.