Materials for Isolation of Nucleic Acids from Whole Blood without a Lysis Step

无需裂解步骤即可从全血中分离核酸的材料

• 批准号: 6990337
• 负责人: Hashem Akhavan-Tafti
• 金额: $ 10万
• 依托单位: LUMIGEN, INC.
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-08-01 至 2006-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6990337
• 关键词: bacterial DNA; biomaterial development /preparation; blood; body fluids; chemical synthesis; gel electrophoresis; human genetic material tag; human tissue; magnetism; method development; nucleic acid purification; nucleic acids; particle; physical separation; virus DNA

DESCRIPTION (provided by applicant): The objective of the proposed study is to develop new materials for the rapid isolation of nucleic acids from complex matrices such as whole blood. In Phase I we will prepare functionalized \microparticles for binding and releasing nucleic acids that utilize proprietary chemistry recently i developed at Lumigen. These materials find many uses but are intended particularly for the rapid ' and simplified isolation of genomic nucleic acids from unpurified or heterogeneous matrices, especially DNA in mammalian tissue or whole blood. This is made possible by a unique feature of these materials: their ability to capture DNA and/or RNA under virtually any reasonable conditions. As a result we have been able to bind DNA from whole blood without any lysis step. The prototype new materials bind nucleic acids with exceptional strength under a range of conditions and resist release under a wide variety of conditions. The unique binding properties of the new materials will be utilized to develop optimized methods for stringently washing and releasing the bound nucleic acid in a highly controllable fashion. Preliminary experiments have demonstrated that a cleavable link between the solid support and the binding moiety can be controllably severed to release nucleic acid back into solution. Nucleic acids purified with first generation materials have been found compatible with downstream applications including amplification technologies such as PCR and LMO, a ligase-based nucleic acid amplification technology invented by the PL We expect to produce one or more general use kits for isolating DNA from whole blood without any lysis step and in a quantity and quality acceptable for use in downstream amplification and diagnostic applications. For the initial phase of the project we expect to develop both non-magnetic and | magnetically responsive microparticles from different solid phase support materials. Since the j prototype new materials have shown exceptionally strong binding to nucleic acids, we plan further to begin exploring the use of the developed materials in capturing nucleic acids from low abundance viral and bacterial infectious agents from samples such as blood and other body fluids. This work serves as a prelude to the development of nucleic acid-based test methods for pathogens in human blood and food products to be undertaken in Phase II.

描述（由申请人提供）： 拟议研究的目的是开发用于从复杂基质（如全血）中快速分离核酸的新材料。在第一阶段，我们将利用我最近在Lumigen开发的专有化学方法制备用于结合和释放核酸的功能化微粒。这些材料有许多用途，但特别是用于从未纯化或异质基质，特别是哺乳动物组织或全血中的DNA中快速和简化地分离基因组核酸。这是由于这些材料的独特功能而成为可能：它们能够在几乎任何合理的条件下捕获DNA和/或RNA。因此，我们已经能够结合来自全血的DNA，而无需任何裂解步骤。原型新材料在一系列条件下以优异的强度结合核酸，并在各种条件下抵抗释放。新材料的独特结合特性将用于开发优化的方法，以高度可控的方式严格洗涤和释放结合的核酸。初步实验已经证明，固体支持物和结合部分之间的可裂解连接可以被可控地切断以将核酸释放回溶液中。已经发现用第一代材料纯化的核酸与下游应用相容，包括扩增技术，如PCR和LMO，PL发明的基于连接酶的核酸扩增技术。我们期望生产一种或多种用于从全血中分离DNA的通用试剂盒，而无需任何裂解步骤，并且其数量和质量可接受用于下游扩增和诊断应用。在项目的初始阶段，我们希望开发非磁性和|来自不同固相载体材料的磁响应微粒。由于原型新材料已经显示出与核酸的特别强的结合，我们计划进一步开始探索所开发的材料在从来自样品如血液和其他体液的低丰度病毒和细菌感染因子中捕获核酸中的用途。这项工作是开发人类血液和食品中病原体核酸检测方法的前奏，将在第二阶段进行。