The stem rust resistance genes Sr24, Sr26, Sr36, and Sr1RSAmigo confer resistance to race TTKSK (= Ug99) of Puccinia graminis f. sp. trit-ici Pers. (Pgt). A collection of 776 cultivars and breeding lines of wheat (Triticum aestivum L.) from all growing regions of the United States were screened with simple sequence repeat and sequence tagged site markers linked to Sr24, Sr26, Sr36, and Sr1RSAmigo to determine frequencies of these genes in U.S. wheat germplasm. Marker efficacy in predicting the presence of these genes was evaluated via comparison with assayed seedling infection type. Among the lines evaluated, the most predominant gene is Sr24, present in hard winter, hard spring, and soft winter wheat lines. Resistance in soft winter wheat is primarily due to Sr36. The 1RS·1AL rye translocation carrying Sr1RSAmigo is present at equal frequencies in hard winter and soft winter wheat. Utilization of marker-assisted selection for stem rust resistance genes can hasten the development of wheat cultivars resistant to TTKSK and its variants and allow for the development of resistance gene pyramids for more durable stem rust resistance.