Over the past 25 years, bacteriophage derived serine integrases have become established as essential tools for synthetic biology. Their simple requirements make serine integrases amenable to a wide variety of applications, from in vitro assembly reactions to complex in vivo DNA rearrangements. The integrase recognizes two simple DNA attachment sites, attB and attP, and recombines them to produce two hybrid sites, attL and attR. The new attachment sites are no longer a substrate for the integrase and, therefore, the reaction is strongly unidirectional. The irreversibility of the integration reaction also means that recombined products are extremely stable, even if the integrase remains present. For some applications the recombination reaction must be reversed, which requires a second protein known as recombination directionality factor (RDF). The RDF binds directly to the C-terminal domain of the integrase to induce a conformational change that allows recombination of attL/attR and inhibits recombination of attB/attP. While Integrases can usually be identified by bioinformatics alone, RDFs are extremely difficult to predict. The few RDFs that are currently known range in size from 6 kDa (SPBc) to 28 kDa (Bxb1), share very little sequence identity with each other and sometimes have secondary biological functions. As increasingly complex applications are developed the requirement for novel integrases and RDFs also increases. I will present our work toward understanding the interactions between the archetypal ϕC31 integrase and RDF and the importance of the 3D structure of the C-terminal domain. We are also working to characterize several newly identified integrases and RDFs from environmental phage, which are now being combined with published integrases to produce bespoke biosynthetic assemblies. Discovery of new RDFs and determining how RDFs control recombination directionality is a vital prerequisite for complex applications such as biocomputing or pathway engineering.