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Abstract The uplift of the Central American Isthmus (CAI) represents a natural laboratory for the study of allopatric speciation in marine organisms. Several geminate species pairs of both vertebrates and invertebrates formed following the uplift of the CAI, including damselfishes of the Pomacentridae family. However, to date no studies have explored the genomic differences among geminate species in the Pacific and Atlantic Oceans. In this study we present genome assemblies for the transisthmian species pair consisting of the Sergeant Major Abudefduf saxatilis (Tropical Atlantic) and the Panamic Sergeant Major Abudefduf troschelii (Tropical Eastern Pacific) derived from PacBio long-read sequencing. The new genomes are near-chromosome level, and among the highest-quality genomes currently available for coral reef fishes. We show that large structural variants distinguish the two species, including a 9 megabase inversion in linkage group (putative chromosome) 6. Additionally, we show through an analysis of demographic history that alleles within the two genomes have different coalescence time distributions, which may be due to different effective population sizes, population structures, and/or selection regimes in the two oceans. Finally, we highlight gene families that were significantly expanded or contracted between A. troschelii and A. saxatilis . Some of these are related to the environmental conditions that differ between the two oceans, such as gamma crystallin M ( crygm ), which is linked to vision, and vitellogenin ( vtg ), which is associated with egg provisioning. These genomes set the stage for comparative analyses of genetic structure and selection on the marine organisms that originated with the formation of the CAI. Significance Statement Understanding speciation in oceans has represented a historical challenge, given the high rates of dispersal during larval stages for most marine groups. There are few geologic events that have propelled marine biodiversity while completely blocking the genetic exchange during the speciation process. The final closure of the Central American Isthmus (CAI), 2.8 million years ago, is one of these few examples where marine populations from the tropical Pacific and Atlantic oceans have been completely isolated, providing the opportunity to study how genomes diverge in the absence of gene flow. Here, we present the first comparative genomics study of marine organisms separated by the CAI, showing that genomes on the two sides of the CAI differ structurally. We also identify contractions and expansions of various gene families between the two species, which are potentially associated with adaptations to the different environmental conditions in the two oceans. This represents a relevant first step for comparative genomics of geminate species isolated by a major geologic event.