DEVELOPMENT OF AN IMPROVED MISMATCH BINDING PROTEIN

描述：（研究者摘要）由于MutS在体内参与DNA错配碱基的识别和修复，因此它可能成为需要在体外定量结合含错配DNA的应用的候选者。然而，溶液中的MutS缺乏特异性，即对含有错配的DNA分子和完美配对的DNA分子的区分能力较差，结合能力较弱。固定化MutS具有良好的识别能力，但容量较低。本项目的目标是从大肠杆菌MutS开始，通过诱变和强大的选择系统，开发一种改进的错配结合蛋白，能够在含有错配或未配对碱基的DNA溶液中紧密结合。这种蛋白质在突变检测、克隆、基因发现和PCR保真度方面具有广泛的用途。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。它可用于当前的Gene Check产品，包括错配结合珠和固定化错配结合蛋白板，并将形成PCR清理试剂盒的基础，可用于突变检测和克隆应用。改进的错配结合蛋白也可能是通过血统基因发现方法进行身份鉴定的基础。

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.