Soyfoods have been consumed for more than 1000 years in Asia, and they are becoming increasingly popular in western diets due to their nutritional value and healthy benefits. Soybean contains high protein and fiber, low carbohydrates, and is also a good source of calcium. The objective of this study was to identify major quantitative trait loci (QTLs) associated with seed calcium content. A mapping population consisting of 179 F2:3 lines was derived from a high calcium x low calcium soybean cross, Camp x SS516. Camp and SS516 are both popular soybean cultivars for the soyfoods market. The HNO3 digestion method was used to extract calcium from soybean seeds and calcium content was determined by ciros spectroscopy. The variation of calcium content in the genetic population followed a normal distribution ranging from 0.19 to 0.42% with an average of 0.29%. The soybean genetic linkage map of calcium content had 148 simple sequence repeats (SSRs) markers, 15 of which were not in the integrated genetic linkage map of the soybean published in 2004. All the markers converted into 19 linkage groups, and covered a genetic distance of 1421.8 cM on soybean genome in the Kosambi function. Based on this linkage map, three QTLs (Ca1, Ca2 , and Ca3 ) on linkage group A2, I, and M were identified by using both single marker analysis (SMA) and composite interval mapping (CIM), and they accounted for 45% of calcium content variation in soybean seeds.