Sesame (Sesamum indicum L.) is one of the oldest and most nutritional oilseed crops, of which domestication history has been poorly understood. This study suggested that sesame has undergone domestication bottleneck during its use for a long time. In this investigation, the molecular analysis included 4.4 Mbp of the genomic DNA of sesame comprising stearoyl acyl desaturase (sad), fatty acid desaturase 2 (fad2) and omega 3 fatty acid desaturase (o3fad) genes in 99 accessions of four populations of sesame germplasm namely: wild species, landraces, improved cultivars and introgressed lines. Results indicated that the improved cultivars and landraces lost 46.6 and 36.7% of nucleotide diversity, respectively, which indicate that the genetic diversity of the crop had been eroded due to selection after domestication. However, there was no significant reduction in genetic diversity of improved cultivars compared with landraces, indicating that unique improved cultivars generated through crosses were of less frequency in this population. Moreover, introgressed lines retained only 17.77% (π) and 4.57% (θ) of landrace diversity. To evaluate the impact of selection across fatty acid biosynthetic pathway, individual nucleotide diversity at three major genes involved in the pathway was surveyed. The analysis between wild and improved cultivars supported positive selection in fad2 and o3fad loci. Though locus-to-locus sequence variation was observed, positive results with two most important loci supported selection after domestication. Reduced diversity in these critical quality governing genes in improved cultivars suggested that future sesame cultivation would benefit from the incorporation of alleles from sesame's wild relatives.