High resolution analysis of linear genomic DNA in Parallel nanochannel arrays

描述（由申请人提供）： 我们正在开发一种纳米芯片设备，用于操纵长基因组DNA，用于癌症生物标志物的高分辨率（DNA酶），全基因组分析，如基因扩增，缺失和易位。这些染色体结构畸变与恶性转化过程密切相关，是许多类型癌症的重要诊断、预后和治疗指标。虽然PCR提供了检测和分析这些异常的最终（单碱基）分辨率，但以全面的线性方式扫描整个基因组是不切实际的。依赖于探测染色体的技术，如中期FISH，虽然提供了一个泛基因组的观点，不能解决低于Mb范围的结构。通过探测未压缩的间期DNA，可以提高分辨率，但基因组的空间组织丢失，因此难以获得多重和定量信息。通过拉伸（线性化）间期DNA，使用诸如“分子梳理”或“光学作图”的技术，可以以空间上显著的方式探测特定的基因座，并且分辨率在kb范围内。然而，用于机械线性化DNA的技术是固有可变的，导致分子的不一致拉伸，其经常交叉并缩回到自身上。这使得它难以标准化，如生物医学界的高通量方法的技术。我们正在开发一种创新的替代机械拉伸DNA的方法。我们已经发现，由于DNA聚合物的自回避性质，单个DNA分子在流入限制性纳米尺度通道（纳米通道）时将以一致的方式伸长和拉直。我们已经使用了一种新的纳米压印光刻技术，可靠地制造硅芯片中的纳米通道结构，并已证明，这些纳米通道中的DNA可以可视化和测量其尺寸。我们现在提出的问题是，我们能否用基因座特异性探针定量地询问这种线性化的DNA，以检测与癌症相关的染色体结构畸变？我们的产品，纳米通道阵列芯片，将包括一个集成平台的一部分，用于DNA结构的常规和标准化定量分析，这将使数据的存档和跨实验室比较成为可能。

DESCRIPTION (provided by applicant): We are developing a nanochip device for manipulating long genomic DNA for high-resolution (kilobase), whole-genome analysis of cancer biomarkers such as gene amplifications, deletions, and translocations. These chromosome structural aberrations are strongly implicated in the process of malignant transformation and are important diagnostic, prognostic, and therapeutic indicators for many types of cancer. Although PCR offers the ultimate (single-base) resolution for detecting and analyzing these anomalies, it is impractical for scanning the entire genome in a comprehensive, linear fashion. Techniques that rely on probing chromosomes, such as metaphase FISH, while providing a pan-genomic view, cannot resolve structures below the Mb range. By probing uncompressed interphase DNA, resolution can be improved, but spatial organization of the genome is lost, so multiplexed and quantitative information is difficult to obtain. By stretching out (linearizing) interphase DNA, using techniques such as "molecular combing" or "optical mapping," it is possible to probe specific loci in a spatially-significant way, and with resolutions in the kb range. However, techniques for mechanically linearizing DNA are inherently variable, leading to inconsistent stretching of molecules, which often cross over and retract upon themselves. This makes it difficult to standardize such techniques as high throughput methods for the biomedical community. We are developing an innovative alternative to mechanical stretching of DNA. We have found that individual DNA molecules, because of the self-avoiding nature of the DNA polymer, will elongate and straighten in a consistent manner when streamed into confining nanometer-scale channels (nanochannels). We have used a novel nanoimprint lithography technique to reliably manufacture nanochannel structures in silicon chips and have demonstrated that DNA in these nanochannels can be visualized and their dimensions measured. We now ask the question, can we quantitatively interrogate this linearized DNA with locus-specific probes for the detection of chromosome structural aberrations associated with cancer? Our product, the nanochannel array chip, will comprise part of an integrated platform for the routine and standardized quantitative analysis of DNA structure that will enable archiving and cross-laboratory comparison of data.