Peganum harmala is a halonitrophilous perennial herb that is relevant in the landscape of steppe, semidesert, and desert territories in southern Europe, northern Africa, and southwestern Asia. We present here data on the nuclear DNA amount assessment in one Iberian and 17 Tunisian populations of this species. The 2C values, belonging to the very small genome category, ranged from 0.61 to 0.67 pg. These data are the first on genome size for the species and for the whole genus and the whole family. In addition, the somatic chromosome number (2n = 24) has been counted in one population, confirming previous reports and constituting the first one in a Tunisian accession of the species. The results show a high degree of homogeneity of the character studied within the species. These new data, filling a gap in genome size knowledge at family level, together with the specific homogeneity in genome size in a large area, could be relevant for further large-reach analyses on genome size evolution.