Point mutations in the ATM gene in malignant B-cell lymphomas with (A, B, C) and without (D) monoallelic deletions of ATM. Mutations are indicated by arrowheads or by a bar. A, B-CLL-A: SSCP analysis of a 242-bp restriction fragment derived from the RT-PCR product containing exons 56-65 identified an aberrantly migrating fragment. DNA sequence analysis of the respective fragment showed 8412delA causing frameshift and truncation of the protein. DNA sequences of PCR amplified exon 59 are shown. (B) B-CLL-B: SSCP analysis of a 307-bp restriction fragment of the RT-PCR product encompassing exons 56-65 and DNA sequence analysis of the corresponding RT-PCR products identified the nonsense mutation 9139C>T (Arg3047ter). Note that the amount of cells with an 11q22-q23 deletion was 58% in the tumor sample, and therefore the signal intensity of the normal allele was unusually high. DNA sequence analysis of exon 65 from patient’s germline DNA showed absence of the mutation in the germ line. (C) MCL-A: The splice-donor site mutation in intron 59 (IVS59+1G>T), which causes skipping of exon 59 from the ATMtranscript and loss of 50 amino acids, was detected by DNA sequencing of the RT-PCR fragment containing exons 56-65 and by sequencing of PCR amplified exon 59 and its flanking intronic regions. The patient’s germ line DNA did not contain the mutation. (D) MCL-B: Cloning and subsequent sequencing of RT-PCR fragments harboring exons 51-57 allowed assignment of the two heterozygous mutations 7268A>G and 7250insGAA to two separate ATM transcripts. Thus, both alleles were affected by two independent mutations.