Root parasitic plants in the Orobanchaceae germinate in response to chemicals exuded by plants, including the hormones strigolactones. Strigolactones are important for beneficial symbioses with fungi and for many aspects of plant development; however, strigolactones are only known to stimulate germination of parasites. Soon after parasitic plants germinate, they can cause irreversible damage to hosts, which often include important crops. Thus, strigolactone-responsive seed germination is a promising target for the development of parasite control strategies. However, little was known about strigolactone perception in parasites until recently.We identified candidate genes for roles in host-responsive seed germination in parasites based on signaling systems in the model non-parasitic plant Arabidopsis thaliana. In A. thaliana, KAI2 responds to smoke-derived germination stimulants called karrikins, and probably an unidentified endogenous signal called KAI2 ligand (KL). Its homolog D14 is required for strigolactone responsiveness in A. thaliana and other model non-parasites. We discovered extensive duplication of KAI2 in parasite genomes. We functionally characterized KAI2 paralogs from parasites by testing them as transgenes in A. thaliana null mutants, and we found that some parasite KAI2 confer strigolactone-responsive seed germination. Other KAI2 paralogs from parasites function more similarly to Arabidopsis KAI2 (AtKAI2), although one responds preferentially to karrikin, and others to KL. Thus, gene duplication and sub- and neofunctionalization have likely shaped KAI2 genes in parasites, some of which enable host-responsive seed germination.We next investigated the molecular basis for differences in likely ligand preference among different KAI2 genes. We modified sites that we identified as potential specificity-determining positions (SDPs) in AtKAI2, and we tested AtKAI2 variants for responsiveness to various signals. Although we did not reconstitute strigolactone responsiveness, we made AtKAI2 more specific for karrikin or KL through different sets of targeted modifications. We also functionally characterized two KAI2 paralogs from lettuce and found that they have likely subfunctionalized relative to AtKAI2, similarly to strigolactone-unresponsive KAI2 from parasites. Together, our results illustrate the complex evolutionary history of KAI2. They also provide valuable information for the fight against parasitic plant infestations, which threaten food security in resource-limited parts of the world.