Creation and manipulation of bipartite expression transgenes in C. elegans using phiC31 recombinase

Abstract

Bipartite expression systems are widely used in model organisms to express specific gene products in a cell-specific context. They are typically encoded as two independent, unlinked transgenic insertions: a driver and a reporter. Herein, I outline an efficient method named recombination-mediated homolog exchange (RMHE) that uses genetically encoded phiC31 recombinase to convert driver and reporter transgenes integrated at the same genetic position from a trans configuration where one component is on each chromosome, to a cis configuration where the driver and reporter are on the same chromosome. This approach is based upon the development of a set of 3’ attB tagged driver lines and 5’ attP tagged reporter lines. This genetic based approach leverages both the power of combinatorial re-assortment of drivers and reporters and the simplicity of single locus genetics. I also describe a novel microinjection-based approach named recombination-mediated integration (RMI) that utilizes the individual attB driver and attP reporter lines as landing site for the phiC31 recombinase mediated integration of whole driver and reporter plasmids into the existing reporter and driver lines, respectively. Thus, this work outlines both a novel genetic based and a novel microinjection-based method to create cis-linked driver/reporter pairs. These new tools increase the utility of bipartite systems for C. elegans genetics by reducing the complexity of reporter system segregation in crosses and thus can greatly simplify the use of bipartite reporter systems during genetic analysis.