Abstract: An efficient genotype-dependent high frequency plant regeneration system was developed for finger millet genotype GN-4 (Gujarat Nagli-4). For callus induction, five different concentrations of 2,4-D (0.5, 1.0, 2.0, 3.0 and 4.0 mg L−1) were used to culture mature seeds and observed notable percentage of callus induction 94.00%; fresh and dry weight (0.903 g and 0.093 g) on Murashige and Skoog (MS) medium containing 2.0 mg L−1 2,4-D. Maximum callus proliferation (92.80%) with good quality of callus were observed on MS basal medium having 2.0 mg L−1 2,4-D + 0.5 mg L−1 BAP + 0.5 mg L−1 kinetin. On transferring callus for shoot generation, most effective regeneration of shoots (88.30%) with higher number of shoots per explant (8.00) and maximum shoot length (6.90 cm) were observed on MS + 1.0 mg L−1 BAP + 1.0 mg L−1 kinetin. For rooting, regenerated shoots were cultured to full-MS and half-MS media enriched with different levels of IBA and NAA and recorded remarkable early rooting response (71.67%) on half-MS medium + 0.1 mg L−1 IBA. These healthy rooted plants were transferred to pots containing sterilized potting mixture consisting vermicompost: soil: coco peat (1:1:1) for hardening and successfully acclimatized in the field conditions. Genetic stability of regenerated plantlets was confirmed by randomly amplified polymorphic DNA (RAPD) with monomorphic bands during different stages of plant tissue culture system and proved no variation. Furthermore, minerals (P, Mg, Ca and Fe) study based on inductively coupled plasma mass spectrometry (ICP-MS) determined higher content of P, Ca and Fe in seeds of regenerated plantlets as compared to mother plant.