REF Select: Expert System Software for Selecting Restriction Endonucleases for Restriction Endonuclease Fingerprinting

Abstract

REF Select, expert system software, has been developed to assist in the selection of optimal restriction endonucleases for restriction endonuclease fingerprinting (REF), a method for rapid and sensitive mutation screening of long DNA segments (1–2 kb). The REF method typically involves six separate digestions with up to two restriction endnonucleases used in each digestion. If done manually, performing a comprehensive review of the large number of possible sets of restriction endonucleases that could be used (over 1019 in the example presented here) and making an optimal choice is not feasible. Furthermore, the typical nonoptimal manual selection takes approximately 8 h by someone experienced with REF. REF Select enables a comprehensive review of the possible sets and a consistent, objective and fast selection of an optimal set by using a two-step strategy: the selection of sets that meet specific constraints, which is followed by a ranking of those sets by an optimality score. Based on our experience with REF, we chose default selection and ranking parameters to help the user get started quickly. These parameters form a knowledge base that can be customized and then saved by the user. In conclusion, REF Select facilitates the general application of REF by serving as an expert system for the selection of optimal restriction endonucleases. We demonstrated REF Select using an example segment from the human p53 gene.