Habitat and seed size are the key factors that affect plant growth and development. The influence of these factors on the life history traits of the wild medicinal Calotropis gigantea is scanty around the globe. Hence, a laboratory and a pot experiment were conducted to investigate the impact of various sources (habitats) of seeds and their sizes on germination, seedling growth, and physiological traits of C. gigantea following a two-factor, completely randomized design with three repetitions. Factor-A comprised five habitats’ seeds (T1-roadside, T2-railway line, T3-river bank, T4-waste dump, and T5-grazing land), and factor-B consisted of three sizes (mass) of seeds (L-large, M-medium, and S-small). In this study, we observed that the grazing land habitats’ seed (T5) performed better than other sources of seeds for various studied traits. The performances of different sizes of seeds were categorized as L > M > S. Overall, it was notable that the greater performances of different habitats’ seeds with larger sizes were characterized as T5 × L > T1 × L > T2 × L > T3 × L > T4 × L for germination, seedling growth, and physiological attributes. The present findings might have provided greater insights into the growth, development, and productivity of C. gigantea useful to farmers, practitioners, and researchers.