The aim of this study was to characterize the collection of cassava (Manihot esculenta) obtained from Côte d’Ivoire based on 14 microsatellite loci. Two hundred (200) accessions from five geographical zones – center, center-west, south, south-west, and west – were evaluated. High diversity values were found among the five above-mentioned regions, with 2.415 ± 0.153 alleles per locus on average. A high percentage of polymorphic loci varying from 38.46–100% was also observed. An average of 0.166 ± 0.026 for observed heterozygosity and 0.242 ± 0.028 for expected heterozygosity were observed too. The mean value of the Shannon index was 0.423 ± 0.048. Most genetic diversity was concentrated within the geographical zone (Hs = 0.236 ± 0.026). A high genetic differentiation among the population was found (Fst = 0.241), indicating the low level of gene flow between the population due to the infrequent sharing of cassava planting materials among farmers between regions. Based on these results, therefore, during the collecting missions, it is recommended to collect enough cassava cutting within each population to have the greatest possible diversity.