Delta-Like Non-Canonical Notch Ligand 1 (DLK1) is one of the key genes involved in the development of muscle, liver, pancreas, and lung cells; adipocytes production; and the improvement of digestion, growth performance, and meat quality. It has been documented that fennel is effective on increasing the DLK1 gene (
DLK1
) expression in the testis, liver, and muscle tissues, which may consequently have important implications for sheep production. Hence, the aim of the current investigation was to evaluate the fennel seed powder's effect on
DLK1
expression in testis, liver, and humeral muscle tissues in growing lambs. For the purpose of this study, 30 male Kermani sheep were fed with three different group of diets (number of animals in each group was 10), including control (without any fennel seed powder), treatment 1 (with 10 g/kg of dry matter (DM) fennel seed powder), and treatment 2 (with 20 g/kg of DM fennel seed powder) during a 3-month period. Thereafter, total RNA was extracted, cDNA was synthesized, and Real-Time PCR was performed. The addition of fennel seed powder (in the treatment 1 and treatment 2 groups) in the growing lambs diets consequently resulted in greater expression of
DLK1
in both the liver and humeral muscle tissues compared to the testis tissue (P < 0.05). Furthermore, the increased
DLK1
expression was higher in the tissue of humeral muscle (P < 0.05) in comparison to the other two tissues. As well, the concentration of blood testosterone was greater (P < 0.05) for the animals fed with fennel powder compared to growing lambs fed with the control diet. However, the concentrations of blood liver enzymes, including serum glutamic oxaloacetic transaminase (SGOT) and serum glutamic pyruvic transaminase (SGPT), decreased by the addition of 10 g/kg DM fennel to diets of lambs compared to the control diet (no fennel). Therefore, it can be concluded that using fennel seed powder in the diet of growing lamb by affecting the expression of
DLK1,
can improve the concentrations of blood testosterone, SGOT, SGPT, and muscle structure (increased mass of muscle and size of muscle fiber).