Site-specific recombination between inverted target sites generates dicentric/acentric chromosomes

Abstract

ABSTRACTSite-specific recombinases are widely used tools for analysis of genetics, development and cell biology, and many schemes have been devised to alter gene expression by recombinase-mediated DNA rearrangements. Because theFRTandloxtarget sites for the commonly used FLP and Cre recombinases are asymmetrical, and must pair in the same direction to recombine, construct design must take into account orientation of the target sites. Both direct and inverted configurations have been used. However, the consequence of recombination between target sites on sister chromatids is frequently overlooked. This is especially consequential with inverted target sites, where exchange between oppositely oriented target sites on sisters will produce dicentric and acentric chromosomes. By using constructs that have inverted target sites inDrosophila melanogasterand in mice, we show here that dicentric chromosomes are produced in the presence of recombinase, and that the frequency of this event is quite high. The negative effects on cell viability and behavior can be significant, and should be considered when using such constructs.