Towards Genomes-to-Design: Building and Testing a Minimal Essential Chromosome

合成生物学遵循工程原理，构建由模块化DNA部分编码的新设备、途径和电路，最近转向活细胞中整个基因组的工程。合成生物学大部分工作的目标是设计和制造细胞，使其发挥在自然界中通常无法发挥的有用新功能。为了进一步推动这一领域的发展，重要的是要设计工作马活细胞，使它们能够可靠地和可复制地执行新任务。为此，人们对开发用最小基因组构建的简化细胞非常感兴趣。通过基因组工程和合成，应该有可能创造出不编码许多基因的基因组，这些基因驱动的过程对于细胞执行其主要功能是不必要的。这里提出的工作旨在加速这种最小基因组的工程和合成，这是第一个在实验室中仅从支持细胞功能的必要元素构建工作染色体的项目。从理论上讲，用这种最小基因组生长的细胞在执行工程任务时应该更有效，比如高产量的药物分子的生物合成。因此，我们的工作将能够产生对生物技术有价值的专业菌株。为了实现这一目标，我们计划利用从我们的工作中获得的知识和工具作为国际Sc2.0项目的一部分，该项目正在构建一个完全合成的面包酵母基因组（Saccharomyces cerevisiae）。我们刚刚为这个项目完成了一条染色体的构建，它现在编码了正常情况下在其天然对应染色体上发现的所有334个基因。在这个项目中，我们的目标是用一个更小的最小版本取代整个染色体，这个版本由DNA模块组成，每个模块编码来自染色体的一个基因，这些基因被认为是实验室细胞生长所必需的。为了做到这一点，我们将使用一种叫做SCRaMbLE的系统，它被硬编码到我们最近完成的合成染色体的DNA中。开启这个系统会导致生长所需的基因自动从生长中的细胞中消失。用我们的工程酵母细胞进行大规模的研究，然后对整个群体进行基因组测序，应该会为我们提供一套丰富的数据，告诉我们哪些基因是酵母在实验室中生长所必需的。有了这个重要的新数据集，我们将继续构建我们的最小合成染色体，并评估其在生长的酵母细胞中取代完整染色体的能力。我们计划测量具有最小染色体的细胞在各种条件下的表现，并确定它们是否能够比正常酵母更高效地生长和表达基因，现在冗余的DNA已经被去除。这将产生重要的新见解，以了解细胞如何有效地消耗资源，并已进化到在其基因组上编码许多非必需基因。这也将使我们有机会生产用于生物技术的新型特殊细胞，我们计划测试我们的最小染色体酵母生产各种药物分子的能力，这些药物分子对工业特别是对英国工业合作者有价值。

Synthetic biology follows engineering principles to build novel devices, pathways and circuits encoded by modular DNA parts, and more recently has turned to the engineering of entire genomes in living cells. The aim of much of the work of synthetic biology is to design and build cells to perform useful new functions they typically don't perform in nature. To further this field, it is important to engineer the workhorse living cell so that they perform their new tasks reliably and reproducibly. To this end there is significant interest in developing simplified cells built with minimal genomes. Through genome engineering and synthesis, it should be possible to create genomes that do not encode the many genes driving processes unnecessary for the cell to perform its main desired function. The work proposed here aims to accelerate the engineering and synthesis of such minimal genomes, by being the first project to build a working chromosome from just the elements deemed essential to support the cell for its function in the lab. Cells growing with this minimal genome should theoretically be more efficient at performing engineered tasks, such as the biosynthesis of a drug molecule at high yields. Our work will therefore be able to produce specialist strains valuable for use in biotechnology.To achieve this, we plan to use knowledge and tools gained from our work as part of the international Sc2.0 project, which is constructing an entirely synthetic genome for baker's yeast (Saccharomyces cerevisiae). We have just completed construction of one chromosome for this project, which now encodes all 334 genes normally found on its natural counterpart. In this project, we aim to replace this entire chromosome with a much smaller minimal version built-up from modules of DNA that each encode one of the genes from this chromosome deemed essential for cell growth in the lab. To do this we will use system called SCRaMbLE that is hard-coded into the DNA of our recently completed synthetic chromosome. Switching this system on leads to genes unnecessary for growth being automatically lost from the growing cells. Doing this at a large scale with our engineered yeast cells and then genome-sequencing whole populations, should provide us with a rich set of data that tells us which genes are required for growth of the yeast in the lab. With this important new dataset in hand, we will then proceed to building our minimal synthetic chromosome and assessing its ability to replace a full chromosome in growing yeast cells. We plan to measure how cells with the minimal chromosome perform in a variety of conditions and determine whether they can grow and express genes with greater efficiency than normal yeast, now that redundant DNA has been removed. This will generate important new insights for understanding how cells consume resources efficiently and have evolved to encode many non-essential genes on their genomes. It will also give us an opportunity to produce new specialist cells for use in biotechnology, and we plan to test our minimal chromosome yeast for their ability to produce a variety of drug molecules that are valuable for industry and particularly for UK industrial collaborators.