Calcium deficiency can lead to osteoporosis. Adequate calcium intake can improve calcium deficiency and prevent osteoporosis. Milk powder is the best source of dietary calcium supplements. Probiotics and prebiotics are considered to be beneficial substances for promoting calcium absorption. In this study, synbiotic milk powder (SMP) was prepared by combining the three, and its calcium supplementation effect and osteogenic activity were evaluated in calcium deficient mice. Through prebiotic screening experiments in vitro, after adding 1.2% iso-malto-oligosaccharide, the number of viable bacteria and the calcium enrichment of Lactobacillus plantarum JJBYG12 increased by 8.15% and 94.53% compared with those of the control group. Long-term calcium deficiency led to a significant reduction in calcium absorption and bone calcium content in mice, accompanied by structural deterioration of bone trabeculae. SMP significantly improved apparent calcium absorption, increased serum calcium and phosphorus levels, and decreased ALP activity and CTX-1 levels. In the meantime, the bone mineral density increased significantly, and the number of bone trabeculae and the proliferation and differentiation of osteoblasts also increased. SMP has good dietary calcium supplementation capacity and bone remodeling ability without significant side effects on major organs. These findings provide insights into using SMP as a dietary calcium source to improve bone health.