Sequence Homology-Independent Protein Recombination (SHIPREC)

Genomic recombination is widely recognized as the principal mechanism by which proteins evolve new functions (1 ). Having realized the importance of recombination in evolution, scientists have developed a variety of methods to mimic this phenomenon in the lab to create libraries of gene chimeras. While the ideal technique would allow for the recombination of any parental genes, most available methods are limited to recombining closely related sequences, i.e., those with ≥70% sequence identity, resulting in crossovers biased towards regions of high identity. This occurs because most recombination methods, e.g., Stemmer-shuffling (2 ), StEP (3 ), RACHITT (4 ), and in vivo methods (5 ), rely on template switching to generate chimeras. However, two methods have recently been described which are capable of generating chimeric libraries independent of sequence identity. Sequence homology-independent protein recombination (SHIPREC) (6 ) is described in this chapter; another method, incremental truncation for the creation of hybrid enzymes (ITCHY) (7 ), is also described in this volume. Both methods are capable of generating chimeric libraries containing all possible single crossovers between the two parental genes.