PCR DETECTION OF XBAL POLYMORPHISM IN THE HUMAN RB GENE OF RETINOBLASTOMA PATIENTS

Inactivation of the Rb (retinoblastoma) tumor suppressor gene is assoc iated Wi and sporadic cases of retinoblastoma and other Rb-related tum ors. Early diagnosis and genetic counseling heavily depend on practica l methods for the detection of Rb deletions and mutations in high-risk families. Here we report on the use of a pair of primers in polymeras e chain reaction (PCR) to amplify a 945-bp fragment from intron 17 of the Rb gene (T.L. McGee, G.S. Cowley, D.W. Yandell and T.P. Dryja, 199 0, Nucleic Acid Research, 18: 207). Xbal digestion of the PCR product reveals 2 allelic versions: a single 945-bp fragment (allele 1) or 2 f ragments of 315 and 630 bp (allele 2). We used total genomic DNA (bloo d and tumors) to investigate the power of this PCR-Rb-Xbal-RFLP in the identification of both segregation and loss of heterozygosity of the Rb gene. In one family studied (family IA) in which 2 generations were affected, it was possible to localize the mutated Rb gene to Xbal-Rb allele 2.The assay of loss of heterozygosity of the Rb gene is availab le for all Xbal-Rb allele 1-2 individuals, so that analyses may be app lied in large scale investigation of the participation of Rb gene in t umor development. We conclude that PCR-Rb-Xbal-RFLP is a practical and powerful tool for oncology research and genetic counseling.