Plantains are a key crop for food security in Africa, particularly in the humid regions of Central and West Africa, where they are a major staple food. Plantain collection of the Centre National de Recherche Agronomique (CNRA) in Côte d’Ivoire contains about 42 accessions, who was little genetically characterized. The objective of this work was therefore to perform molecular characterization of this collection and assess relationship among known genotypic groups (populations) using Simple Sequence Repeat (SSR) loci. The microsatellite profiles of 42 plantain genotypes, were detected a total of 14 alleles, with an average of 2.33 alleles per locus. A high level of genetic variation was obtained within accessions, with the Shannon diversity index (I) and the Nei genetic diversity index (He) respectively of I = 0.726 ± 0.023 and He = 0.506 ± 0.008. The mean values of observed heterozygosity (Ho = 0.897 ± 0.025) were higher than the expected mean heterozygosity values (He = 0.506 ± 0.008), which confirms plantain’s highly heterozygous nature. The study also revealed that the SSR loci used, were not able to split the plantains into their known distinct morpho groups based on inflorescence characteristics (French plantain, True Horn and False Horn). The result of this work will contribute to the conservation strategies.