Hunting Rare Human Variations using DNA Sudoku

描述（由申请人提供）：在过去的几年里，我们经历了人类遗传学的范式转变。越来越多的证据强调了罕见的遗传变异在各种各样的性状和疾病中的关键作用。研究罕见的变异是大海捞针的问题，因为为了捕获变异并获得统计能力，必须对大量群体进行测序。高通量测序的性能呈指数级增长，提供了足够的能力来分析大量的标本。然而，样品制备方案不能随测序能力而扩展。准备百分之一到几千个标本进行测序的蛮力方法既麻烦又不划算。因此，下一个挑战是开发一种可扩展的技术，以绕过样品制备中的瓶颈。我们最近设计了一种测序策略，叫做DNA数独，它有巨大的潜力来解决这一挑战。DNA数独是基于根据组合模式汇集样本作为多路复用的一种手段，这大大降低了大规模实验的成本。我们已经证明了DNA数独的可行性，通过分析数以万计的细菌菌落在单次测序运行。我们建议利用我们的概念验证研究的成功，并将DNA数独应用于大型人类群体中罕见遗传变异的研究。作为一个测试案例，我们计划使用该策略通过对1000个种族匹配个体的队列进行风险位点测序来研究与犹太遗传疾病有关的罕见变异。这种前瞻性的方法将首次创建一个全面的风险等位基因目录，并可以立即具有临床意义。拟议项目的成功有可能产生深远的技术影响。它将为各种单基因和复杂疾病的研究引入一种极具成本效益的测序策略，克服基因组学的一个关键障碍。DNA测序的未来发展将进一步增加对可扩展复用方案的需求，使我们的策略具有丰富的理论基础和多种应用。

DESCRIPTION (provided by applicant): In the past few years, we have experienced a paradigm shift in human genetics. Accumulating lines of evidence have highlighted the pivotal role of rare genetic variations in a wide variety of traits and diseases. Studying rare variations is a needle in a haystack problem, as large cohorts have to be sequenced in order to trap the variations and gain statistical power. The performance of high throughput sequencing is exponentially growing, providing sufficient capacity to profile an extensive number of specimens. However, sample preparation schemes do not scale as sequencing capacity. A brute force approach of preparing hundredths to thousands of specimens for sequencing is cumbersome and cost-prohibited. The next challenge, therefore, is to develop a scalable technique that circumvents the bottleneck in sample preparation. We have recently devised a sequencing strategy, called DNA Sudoku, which has a tremendous potential to address this challenge. DNA Sudoku is based on pooling the specimens according to combinatorial patterns as a means of multiplexing, which substantially reduces costs of large scale experiments. We have demonstrated the feasibility of DNA Sudoku by profiling tens of thousands of bacterial colonies in a single sequencing run. We propose to leverage the success of our proof-of-concept study and to adapt DNA Sudoku for the study of rare genetic variations in large human cohorts. As a test case, we plan to use the strategy to study rare variations that are implicated in Jewish genetic diseases by sequencing risk loci of a cohort of 1000 ethnically matched individuals. This proactive approach will create, for the first time, a comprehensive catalogue of risk alleles and can have immediate clinical implications. Success of the proposed project has the potential for a far-reaching technological impact. It will introduce an ultra cost-effective sequencing strategy for a wide-variety of studies on monogenic and complex diseases, overcoming a critical barrier in genomics. Future developments in DNA sequencing will further increase the requirement for scalable multiplexing schemes, rendering our strategy a long lasting one, with rich theoretical foundations and plurality of applications. PUBLIC HEALTH RELEVANCE: Finding individuals that carry disease alleles in their genome is a needle in a haystack problem. As a solution, we propose to develop an innovative and highly efficient DNA sequencing strategy that is based on recent breakthroughs in the field of signal processing and is reminiscent of solving Sudoku puzzles. The strategy will be used to address a long standing problem - creating a comprehensive catalogue of risk alleles for severe genetic diseases, which has immediate clinical implications and can prevent devastating cases.