A microsatellite fluorescent method for linkage analysis in familial retinoblastoma and deletion detection at the RBI locus in retinoblastoma and osteosarcoma

Linkage analysis at the retinoblastoma locus (RB1) is essential for identif ying individuals at risk and to offer adequate genetic counseling in famili al retinoblastoma. It can also be used to detect large deletions involving RE1, which accounts for 15% of the genetic alterations in hereditary retino blastoma. These studies are usually carried out with lengthy Southern blot analyses of relatively uninformative restriction fragment length polymorphi sms. The authors report an alternative, reliable protocol for genotyping th e RE1 locus using two pairs of highly informative intragenic and flanking m icrosatellites linked closely to the RB1 gene, and analysis of the fluoresc ent-labeled polymerase chain reaction products with automatic sizing techno logy. This methodology has successfully identified high risk carriers in fi ve of the five pedigrees of familial retinoblastoma studied. In addition, g ross deletions affecting the RB1 gene were identified in two of 12 sporadic bilateral retinoblastomas, and loss of heterozygosity at the RE1 locus has been detected in one of three osteosarcomas using the same experimental pr otocol. The described protocol is simpler and faster than conventional Sout hern blot methodologies and can identify a larger number of informative cas es.