Eight-day-old seedlings of seven soft red winter wheat cultivars and four elite breeding lines were tested for the presence of powdery mildew resistance genes with 14 isolates of Blumeria graminis f. sp. tritici. These isolates were characterized for their virulence on differential wheat cultivars and lines with single genes for powdery mildew resistance. Phenotypic reactions of the cultivars and lines to each isolate were assessed 8 days after inoculation. Differential reactions and pedigree information were used to determine the presence of putative powdery mildew resistance genes. The B. g. tritici isolates detected powdery mildew resistance gene Pm3a in AGRA brand GR915, and pedigree information indicated that Freedom contained Pm8. No Pm genes were detected in Cardinal, Clark, Dynasty, or Titan. The gene Pm3a was present in breeding lines OH470 and OH493-1, and gene Pm17 was detected in OH464. OH490 contained gene Pm2 or Pm6, or possibly both; the isolates could not differentiate these two genes.