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Romantic relationship satisfaction (RRS) is important for mental/physical health but varies greatly across individuals. To date, we have known little about the biological (genetic and neural) correlates of RRS. We tested the hypothesis that the serotonin transporter promoter polymorphism (5-HTTLPR), the promoter region of the gene SLC6A4 that codes for the serotonin transporter protein, is associated with individuals’ RRS. Moreover, we investigated neural activity that mediates 5-HTTLPR association with RRS by scanning short-short (s/s) and long-long (l/l) homozygotes of 5-HTTLPR, using functional MRI, during a Cyberball game that resulted in social exclusion. l/l compared with s/s allele carriers reported higher RRS but lower social interaction anxiety. l/l compared with s/s carriers showed stronger activity in the right ventral prefrontal cortex (RVPFC) and stronger functional connectivity between the dorsal and rostral ACC when being excluded from the Cyberball game. Moreover, the 5-HTTLPR association with RRS was mediated by the RVPFC activity and the 5-HTTLPR association with social interaction anxiety was mediated by both the dorsal–rostral ACC connectivity and RVPFC activity. Our findings suggest that 5-HTTLPR is associated with satisfaction of one’s own romantic relationships and this association is mediated by the neural activity in the brain region related to emotion regulation.