A Strategy for High Quality Clinical Resequencing of the Human Genome

描述(由申请人提供)：这项建议的总体目标是开发一种策略，以实现人类基因组的临床重新测序，我们将其定义为将人类基因组重新测序应用于大量临床人群，用于疾病基因发现研究以及具有诊断潜力的临床应用。我们的战略依赖于太平洋生物科学平台所使用的“第三代”单分子DNA测序。我们预计，单分子DNA测序技术最终将成为测序器的选择，因为这些系统可以从患者样本中快速收集大量带有临床结果注释的DNA序列，并且成本效益高，操作成本仅为其他测序技术目前成本的一小部分。这第一个目标侧重于开发我们所称的间歇性分段重新排序方法。一组散布的读数将从单个DNA分子中产生，其工作方式与配对序列基本相同，但具有内部间隔序列，而不是通过配对末端方法产生的末端序列对。第二个目标涉及测试基于溶液的方法，以与基因组鸟枪测序相结合，大幅增加临床上高度感兴趣的基因组学区域的折叠复盖率。考虑到它们的随机序列分布，鸟枪式基因组测序努力平等地对待基因组的所有区域。与基因组的其余部分相比，有选择地增加这些感兴趣区域的折叠覆盖率将改善对重要临床变异的检测。 与公共卫生相关：为每个人创建一个“个性化”的药物策略需要破译一个人的癌症的DNA序列，这包括识别任何患者的基因变异，并发现这些变异的临床意义。这些变异的组成和谱可能因个体而异，因此指出识别所有具有临床后果的关键变异的重要性。新的测序技术为创建个性化的基因签名开辟了道路，这些签名可用于定制个人的临床管理，但考虑到数以千计患有特定疾病的人的基因组已经测序，还需要更具成本效益的方法。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to develop a strategy towards clinical resequencing of the human genome which we define as the application of human genome resequencing to large clinical populations for disease gene discovery research as well as a clinical application that has diagnostic potential. Our strategy relies on "third generation" single molecule DNA sequencing as utilized by the Pacific Biosciences platform. We anticipate that single molecule DNA sequencing technology will ultimately be the sequencer of choice given these systems' rapid collection of massive amounts of DNA sequences from patient samples with clinical outcome annotation, and cost-effective operation at a fraction of the current costs of other sequencing technologies. This first goal focuses on developing what we term an intermittent segment resequencing approach. A set of interspersed reads will be generated from a single DNA molecule, much the same way as mate paired sequence works, but having internal interval sequences rather than a terminal sequence pair as generated by paired-end approaches. The second goal involves testing in-solution based approaches to increase substantially the fold coverage of genomics regions of high clinical interest in combination with genome shotgun sequencing. Given their random distribution of sequences, shotgun genome sequencing efforts treat all regions of the genome equally. Increasing the fold-coverage of these regions of interest selectively, in comparison to the rest of the genome, would improve the detection of important clinical variants. PUBLIC HEALTH RELEVANCE: Creating a "personalized" medicine strategy for each individual requires decoding the DNA sequence of a person's cancer that involves identifying genetic variations for any patient and discovering the clinical significance of these variants. The composition and spectrum of these variants can vary significantly from individual to individual, thus pointing out the importance of identifying all of the critical ones with clinical ramifications. New sequencing technologies have opened the way for creating personalized genetic signatures which can be used to tailor the clinical management of individuals but even more cost-effective approaches are needed given that thousands of individuals with specific disease have their genomes sequenced.