Electronically Addressable DNA Assembly

常规地合成整个基因组的能力是人类剩余的巨大技术抱负之一 合成生物学革命。现有组件使用的退火反应设计的复杂性 工作流程是基因组规模DNA生产的根本障碍。有一个实际的数量限制 独特的序列，可以在单一步骤中可靠地自组装成所需的结构，并且是传统的 组装反应的规模通常限制在数十或数百个寡核苷酸。电流并联 合成技术可以产生超过100万个寡核苷酸的池，这对于一个 基因组规模的构建，但远比在单一反应中可靠地组装要复杂得多。尽管 在提高寡核苷酸合成产量方面有很大兴趣，这种技术不能被利用 完全没有配套的引导他们集会的手段。 我们已经取得了进展，能够以100倍于任何已发表系统的产量产生寡核苷酸。这个 拟议的研究工作将这种进步(高达100倍的增强)转化为基因组装的吞吐量。 其基本概念是在能够控制细胞定位的底物上进行DNA合成 寡核苷酸在随后的组装过程中。如果成功，这种方法将解决一个根本的问题 寡核苷酸合成和组装之间的可扩展性不匹配，使新一代基因成为可能 一次合成就能超过人类基因组规模的合成技术。这 项目可能导致100倍的并行化增加和100倍的成本降低，并可能增加汇编 精确度。

The ability to routinely synthesize entire genomes is one of the great remaining technical aspirations of the synthetic biology revolution. The complexity of designing the annealing reactions used by existing assembly workflows is a fundamental barrier to genome-scale DNA production. There is a practical limit to the number of distinct sequences which can reliably self-assemble into the desired construct in a single step, and traditional assembly reactions are typically limited in scale to tens or hundreds of oligonucleotides. Current parallel synthesis technologies can generate pools of over 1 million oligonucleotides, which is entirely inadequate for a genome-scale construction, yet far more complex than can be reliably assembled in a single reaction. Despite substantial interest in improving oligonucleotide synthesis throughput, such technologies cannot be leveraged fully without accompanying means of guiding their assembly. We have made advancements to generate oligonucleotides at a throughput 100× any published system. The proposed studies work to translate this advance (up to 100× enhancement) to the throughput of gene assembly. The underlying concept is to perform DNA synthesis on a substrate capable of controlling the localization of the oligonucleotides during their subsequent assembly. If successful, the approach would address a fundamental scalability mismatch between oligonucleotide synthesis and assembly, enabling a new generation of gene synthesis technologies capable of exceeding the scale of the human genome with a single synthesis run. This project may lead to a 100x parallelization increase and 100x cost decrease, and potentially increase assembly accuracy.