Mutation detection and genetic counseling in retinoblastoma using heteroduplex analysis

Gene diagnosis is essential for confident presymptomatic prediction, genetic counseling, and early management of hereditary retinoblastoma. In screening the leukocyte DNA of three patients with bilateral retinoblastoma for RB1-gene heterozygous germline mutations, we identified mutations involving exon 3 or 18 of the RB1 gene by using heteroduplex analysis and sequencing. In one case the mutation was a 2 bp GT deletion resulting in the loss of the exon 18 splicing-donor; another mutation was a G-to-T transversion at codon 580 in exon 18, which converts Arg to a stop codon. The third mutation involved in 1 bp deletion at codon 96 in exon 3, which leads to a premature stop codon at codon 110. We used information from this heteroduplex technique for genetic counseling and presymptomatic prediction. A newborn was identified as normal, using gene diagnosis; his 15-month follow-up confirmed our prediction.