Watermelon (Citrullus lanatus) seed is rich in oils and the fatty acid composition of watermelon seeds is a significant source of nutrition and income for especially Africa, the Middle East, and Asia countries. Although the fatty acid profile of watermelon seed is well documented, the underlying associations with seed traits have not yet been studied. Therefore, the current study aimed to determine both fatty acid profile of watermelon seeds and the relations between the fatty acid profile and seed/seedling characteristics. For this purpose, two common watermelon cultivars (Alaska F1 and Galaktika) of seed were used to examine the first count, germination percentage, vigour, vigour index, seed coat ratio, 100-seed weight as well as fatty acid profile. Vegetative traits including seedling height, diameter, dry matter, and leaf area were also determined in these cultivars. As a result, there were significant differences among cultivars for vigour, vigour index, seedling height, seedling diameter, and fatty acids. In addition, linoleic (C18:2n-6) (68%), oleic (C18:1n-9) (15%), and palmitic (C16:0) acids (9.7%) were considerably highest in concentration in the watermelon seeds. The correlation analysis showed that palmitoleic acid (C16:1n7) and arachidic acid (C20:0) in watermelon seeds could play an important role in signalling seed and seedling traits. Also, seedling dry matter can be used indicator of a large number of fatty acids based on the correlation results. According to principal component analyses, among several fatty acids, C16:1n-7, C18:1n-9, and monounsaturated fatty acid (MUFA) were found close to each other. The path analyses indicated that MUFA, C16:1n-7, and C18:1n-9 could be used as markers to estimate the first count, germination rate, and vigour of watermelon seeds. To the best of our knowledge, this is the first research that aimed to determine seed and seedling traits of watermelon cultivars based on the fatty acid profile