Cultivated sorghum [Sorghum bicolor (L.) Moench] is an important food security crop in the semi-arid regions of the world including Asia and Africa. Its genetic diversity is contained mostly in traditional varieties and modern cultivars used by farmers. In this study, agro-morphological traits and molecular markers were used to assess genetic diversity in 22 accessions of cultivated sorghum from five countries (Botswana, Namibia, Swaziland, Zambia and Zimbabwe) in the Southern African Development Community (SADC) region. The study revealed a significant variation among 22 accessions in both qualitative and quantitative morphological traits, indicating the accessions’ promising potential as breeding material. For molecular analysis, 11 microsatellite primer-pairs were used, and generated a total of 70 alleles across 20 accessions. Analysis of molecular variance revealed a high level of genetic variation; 67 % among the accessions and 10 % among the five countries. The patterns of genetic diversity and the relationships observed in this study should provide insights for genetic resource conservation and utilization of sorghum germplasm in the SADC region.