DNA HYBRIDIZATION ENHANCEMENT THROUGH DNA BENDING

描述：（改编自申请人的摘要）最近，Steitz和 他的同事们提出，蛋白质诱导的弯曲有助于促进 DNA聚合酶的DNA校对过程。 申请人 重新解释这一观察结果，假设DNA碱基配对 弯曲的DNA双链体的选择性实际上更大 而不是线性（无应力）形式。 他们建议设计和减少 实践DNA杂交分析的化学方法， 模拟了通过弯曲增强选择性的自然过程。 他们 建议设计和合成修饰的核酸探针， 带有一对侧链，在结合它们的同源DNA或RNA后， 目标和交联的侧链，将捕获双链体， 其弯曲的螺旋形状。 在两步结合试验中，将允许探针 以结合到靶修饰的微量滴定表面，然后 化学交联步骤以连接探针侧链对。 交联导致探针连接到表面（通过交联剂）。 目标）作为假结，因此赋予对极端 洗涤条件。 以这种方式，标记的探针结合到表面 仅当双链体形成 并且发生了探针介导的交联。 根据拟议的 工作，第二阶段的应用程序将制定发展这一 一套核酸杂交的工具 增强，特别是作为人类DNA微阵列技术的一部分， 诊断和基于人群的筛查。 拟议商业应用：不可用

DESCRIPTION: (adapted from applicant's abstract) Recently, Steitz and colleagues have proposed that protein-induced bending helps facilitate the DNA proofreading process by DNA polymerases. The applicants reinterpret that observation to hypothesize that DNA base pairing selectivity is actually greater out of the bent form of a DNA duplex than in its linear (unstressed) form. They propose to design and reduce to practice a chemical approach to DNA hybridization analysis which mimics the natural process of selectivity enhancement by bending. They propose to design and synthesize nucleic acid probes which are modified with a pair of side chains which, after binding their cognate DNA or RNA target and crosslinking of the side chains, will trap the duplex into its bent helix form. In a two step binding assay, probe will be allowed to bind to the target-modified microtitre surface, followed by a chemical crosslinking step to link the probe side chain pair. Crosslinking causes the probe to be linked to the surface (through the target) as a pseudoknot and therefore confers resistance to extreme washing conditions. In that way, labeled probe binding to the surface will be detected after stringent washing only if both duplex formation and probe-mediated crosslinking had occurred. Based upon the proposed work, a Phase II application will be formulated to develop this chemistry into a set of tools for nucleic acid hybridization enhancement, especially as part of DNA microarray technology for human diagnostics and population based screening. PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE