Diagnosis of fragile X syndrome by direct mutation analysis

A total of 27 fragile X pedigrees consisting of over 100 nuclear families were analyzed by Southern blotting methods and probes StB12.3 and StB12.3xx to detect the expansion of the (CGG)n repeat within the FMR-1 gene and the abnormal methylation pattern of the adjacent DNA region responsible for the fragile X syndrome. Clinical expression was found to be associated with the presence of a full mutation (delta > 500 bp, associated with abnormal methylation) in all the males and 50% of the females studied, whereas individuals carrying a premutation (delta = 100-700 bp) were normal. A preferential size increase in the enlarged (CGG)n repeat was detected in successive generations, the instability being stronger when transmitted from a female than from a male. No expansion of the premutation to the full mutation occurred in the paternal transmissions, and the size increase was significantly smaller than in the maternal transmissions. This could partly explain the stability of the premutation through several generations in families with transmitting males. In the maternal transmissions, the risk of expansion of a premutation to a full mutation appeared to depend on its size. The critical maternal premutation size leading invariably to the full mutation was between delta = 175-200 bp. This is important for genetic counseling and also explains the commonly observed clustering of affected individuals in fragile X families.