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

合成生物学遵循工程原理，建立由模块化DNA部分编码的新设备、途径和电路，最近转向在活细胞中设计整个基因组。合成生物学的大部分工作的目的是设计和建造细胞，以执行它们在自然界中通常不具备的有用的新功能。为了进一步发展这一领域，重要的是设计主力活细胞，使它们能够可靠地和可重复地执行它们的新任务。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，应该可以创造出不编码许多基因的基因组，这些基因驱动着细胞执行其主要预期功能所不必要的过程。这里提出的工作旨在加速这种最小基因组的设计和合成，这是第一个仅利用被认为对支持细胞在实验室中发挥功能所必需的元素来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率，比如以高产量合成药物分子。因此，我们的工作将能够生产出对生物技术有价值的专业菌株。为了实现这一目标，我们计划将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，该项目正在为面包酵母(Saccharmyces Cerevisiae)构建一个完全合成的基因组。我们刚刚完成了这个项目的一条染色体的构建，现在它编码了正常情况下在自然对应物上发现的所有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来取代整个染色体，这个版本是由DNA模块组成的，每个DNA模块编码这个染色体中被认为对实验室细胞生长至关重要的一个基因。为了做到这一点，我们将使用一种称为加扰的系统，这种系统被硬编码到我们最近完成的合成染色体的DNA中。启动这一系统会导致生长所必需的基因自动从生长中的细胞中丢失。用我们的工程酵母细胞进行大规模的这项工作，然后对整个种群进行基因组测序，应该会为我们提供一组丰富的数据，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，并评估其在生长中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，并确定它们是否可以比正常酵母更高效地生长和表达基因，因为多余的DNA已经被去除。这将为理解细胞如何有效地消耗资源以及如何进化到在其基因组上编码许多非必要基因产生重要的新见解。这也将使我们有机会生产用于生物技术的新的专业细胞，我们计划测试我们的最小染色体酵母生产各种对工业，特别是对英国工业合作伙伴有价值的药物分子的能力。

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.