Polony Sequencing and the $1000 Genome

描述（由申请人提供）：我们建议进一步开发和利用超高通量聚合酶克隆基因组测序，主要目标是在大约一周内（包括文库制备和测序）生成原始数据以重新测序人类基因组，费用低于1，000美元。 目前，这项技术已经非常先进，但还需要进一步的进步才能实现我们的目标。 作为该项目的关键定量里程碑，我们将报告人类基因组的序列，其“靶序列质量等于或优于2002年12月发表的小鼠组装体的靶序列质量（Nature 420：520，2002）”。 该项目分为四个具体目标，即（1）使用涉及酶切的循环连接策略增加polony测序读段长度，（2）改进文库构建和乳化方案，（3）通过移动到替代克隆扩增策略来增加读段密度（除了乳液PCR或ePCR），和（4）扩展我们的软件能力以允许来自我们新提出的测序原始数据的SNP调用。 实现我们目标的进展已进入后期阶段。 我们能够对细菌基因组进行常规测序，我们即将对整个人类基因组进行测序，达到所需的质量水平。 总的来说，我们认为，通过追求本文所述的目标，可以获得大量回报。 我们在拟议的技术方面拥有丰富的经验，我们有一条通往1,000美元基因组的明确道路。 公共卫生相关性：在此，我们建议进一步开发和利用Polony测序技术，其主要目标是在大约一周内以低于1000美元的成本对人类基因组进行测序。 我们提出了几种方法，将逐步推动我们走向和超越1,000美元的人类基因组。 我们坚信，正如所提出的那样，polony测序技术可以实现1,000美元的基因组目标，无疑会彻底改变医学。

DESCRIPTION (provided by applicant): We propose to further develop and utilize ultra-high throughput polony genome sequencing with the primary goal of generating raw data to re-sequence the human genome in about one week (including library prep and sequencing) for less than $1,000. Currently, the technology is well advanced, but further progress is needed to meet our goals. As the critical quantitative milestone of the project, we will report the sequence for a human genome with "a target sequence quality equivalent to or better than that of the mouse assembly published in December 2002 (Nature 420:520, 2002)". The project is divided into four specific aims, which are to (1) increase the polony sequencing read length using a cyclic ligation strategy that involves enzymatic cleavage, (2) improve the library construction and emulsion protocols, (3) increase read density by moving to alternative clonal amplification strategies (other than emulsion PCR or ePCR), and (4) extend our software capabilities to allow SNP calls from our new proposed sequencing raw data. Progress towards our goals is at an advanced stage. We are able to routinely sequence bacterial genomes and we are on the verge of sequencing an entire human genome to the required quality level. Overall, we feel that there are substantial rewards to be gained by pursuing the goals described herein. We have extensive experience with the proposed technologies and we have a clear path toward the $1,000 genome. PUBLIC HEALTH RELEVANCE: Herein we propose to further develop and utilize Polony Sequencing Technology with the primary goal of sequencing the human genome in about one week at a cost of less than $1,000. We propose several approaches that will progressively move us toward and beyond the $1,000 human genome. We firmly believe that polony sequencing technology, as proposed, can achieve the $1,000 genome goal and undoubtedly revolutionize medicine.