DEVELOPMENT OF AN IMPROVED MISMATCH BINDING PROTEIN

改进的错配结合蛋白的开发

• 批准号: 2866674
• 负责人: ROBERT E WAGNER
• 金额: $ 8.72万
• 依托单位: GENE CHECK, INC.
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2000-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2866674
• 关键词: DNA binding protein; bacterial proteins; binding proteins; gene mutation; polymerase chain reaction; protein binding; protein engineering; site directed mutagenesis

DESCRIPTION: (Investigator's abstract) MutS would appear, because of its involvement in vivo in the recognition and repair of mismatched bases in DNA, to be a likely candidate for applications requiring quantitative binding of mismatch containing DNA in vitro. However, MutS in solution suffers from lack of specificity, i.e., poor discrimination between mismatch containing and perfectly paired DNA molecules and from rather weak binding. Immobilized MutS has excellent discrimination, but low capacity. It is the aim of this project to start with E.Coli MutS and, by utilizing mutagenesis and a powerful selection system, develop an improved mismatch binding protein capable of extremely tight binding in solution to DNA containing mispaired or unpaired bases. Such a protein would have widespread utility in mutation detection, cloning, gene discovery and PCR fidelity. PROPOSED COMMERCIAL APPLICATION: An improved mismatch binding protein will have immediate and widespread commercial applications. It may be used in current Gene Check products including mismatch binding beads and immobilized mismatch binding protein plates and will form the basis of a PCR clean up kit, which can be used in both mutation detection and cloning applications. Improved mismatch binding protein may also be the basis of an identity by descent gene discovery method.

产品说明：由于MutS在体内参与DNA中错配碱基的识别和修复，因此它似乎是需要体外定量结合含有错配的DNA的应用的可能候选者。 然而，溶液中的MutS缺乏特异性，即，含有错配的DNA分子和完全配对的DNA分子之间的差的区分以及相当弱的结合。固定化的MutS具有优异的区分性，但容量低。本项目的目的是从大肠杆菌MutS开始，通过利用诱变和强大的选择系统，开发一种改进的错配结合蛋白，该蛋白能够在溶液中与含有错配或未配对碱基的DNA非常紧密地结合。这样的蛋白质将在突变检测、克隆、基因发现和PCR保真度中具有广泛的用途。建议的商业应用：一种改进的错配结合蛋白将有直接和广泛的商业应用。它可以用于当前的基因检查产品，包括错配结合珠和固定的错配结合蛋白板，并将形成PCR清除试剂盒的基础，该试剂盒可用于突变检测和克隆应用。改进的错配结合蛋白也可以是通过血统基因发现方法的同一性的基础。