An effective mutation-based measure for evaluating the suitability of parental sequences to undergo DNA shuffling experiments

The DNA shuffling process has been successfully used in many experiments of Directed Molecular Evolution. In a shuffling experiment genes are recombined by an iterative procedure of PCR cycles aiming at obtaining new genes, hopefully with some of the original functions being improved. The optimizations of the parameters involved in the process as well as the characteristics of the parental sequences are of extreme importance to guarantee the success of a shuffling experiment. This paper proposes a new measure, based on the number of bases between existing mutations in the parental sequences, suitable for evaluating the suitability of two sequences to be submitted to a DNA shuffling experiment. In order to investigate the usefulness of the proposed mutation-based measure versus two commonly used measures, a family of 37 DNA gene sequences codifying for snake venom metallopeptidases was used for evaluation purposes using the three measures. The parental sequences identified by each of the three measures were validated by simulating the DNA shuffling process using the software eShuffle. The eShuffle results illustrate on the benefits of the mutation-based measure proposed in this paper.