Detection of heterozygous mutations in the RB1 gene in retinoblastoma patients using single-strand conformation polymorphism analysis and polymerase chain reaction sequencing

Abstract

Several families segregating the autosomal dominant form of the hereditary retinoblastoma predisposition gene have been analysed for the causative mutation. We have used the single-strand conformation polymorphism (SSCP) technique to screen for mutations, exon by exon, in the RB1 gene in affected patients from these families. The SSCP technique has proved a rapid and simple technique which relies on the sequence-dependent migration of single-stranded DNA in a non-denaturing polyacrylamide gel. Oligonucleotide primers flanking all 27 exons and the promoter region of the RB1 gene are reported here. The polymerase chain reaction (PCR)-amplified products range in size from 212 to 625 bp and incl..