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.
