Screening of AML1 mutations by a nonisotopic RNase cleavage assay (NIRCA). (A) Structure of the AML1 protein and strategy of RT-PCR. The primers for PCR are shown by small directed arrows below the diagrammatic structure of AML1. The dotted lines with double arrowheads indicate the segments amplified by RT-PCR. Mutations found in 8 patients are mapped onto the segment containing the Runt domain: (○) silent; (•) missense; (□) frameshift; (▪) nonsense. (B) Gel electrophoretic patterns of RNAs generated by NIRCA. The sense strand from each test sample was hybridized with the antisense strand from the wild-type in the upper panel and vice versa in the lower panel. Lanes 1 through 8, patients numbered respectively; lanes 9 and 10, wild-type controls. M denotes Hpa II-digested pUC19 DNA as size markers. The arrowheads on the right indicate the position of original RNA duplexes. For patients no. 4 through 6, cleavage products were recognizable only in the upper panel.