MUTATION DETECTION IN MACHADO-JOSEPH DISEASE USING REPEAT EXPANSION DETECTION

Background: Several neurological disorders have recently been explaine d through the discovery of expanded DNA repeat sequences. Among these is Machado-Joseph disease, one of the most common spinocerebellar atax ias (MJD/SCA3), caused by a CAG repeal expansion on chromosome 14. A u seful way of detecting repeat sequence mutations is offered by the rep eat expansion detection method (RED), in which a thermostable ligase i s used to detect repeat expansions directly from genomic DNA. We have used RED to detect CAG expansions in families with either MJD/SCA3 or with previously uncharacterized spinocerebellar ataxia (SCA). Material s and Methods: Five MJD/SCA3 families and one SCA family where linkage to SCA1-5 had been excluded were analyzed by RED and polymerase chain reaction (PCR). Results: An expansion represented by RED products of 180-270 bp segregated with MJD/SCA3 (p < 0.00001) in five families (n = 60) and PCR products corresponding to 66-80 repeat copies were obser ved in all affected individuals. We also detected a 210-bp RED product segregating with disease (p < 0.01) in a non-SCA1-5 family (n = 16), suggesting involvement of a CAG expansion in the pathophysiology. PCR analysis subsequently revealed an elongated MJD/SCA3 allele in all aff ected family members. Conclusions: RED products detected in Machado-Jo seph disease families correlated with elongated PCR products at the MJ D/SCA3 locus. We demonstrate the added usefulness of RED in detecting repeat expansions in disorders where linkage is complicated by phenoty ping problems in gradually developing adult-onset disorders, as in the non-SCA1-5 family examined. The RED method is informative without any knowledge of nanking sequences. This is particularly useful when stud ying diseases where the mutated gene is unknown. We conclude that RED is a reliable method for analyzing expanded repeat sequences in the ge nome.