Improvement in the molecular diagnosis of Machado-Joseph disease

Background: Direct detection of the gene mutation allows for the confirmati on of the clinical diagnosis of Machado-Joseph disease (MJD), the most freq uent cause of autosomal dominant spinocerebellar ataxia worldwide. Objective: To address the main difficulties in our national MJD predictive testing program. The first was the emergence of intermediate alleles, for w hich it is not yet possible to determine whether they will cause disease. T he second was the issue of homoallelism, ie, homozygosity for 2 normal alle les with exactly the same (CAG)(n) length, which occurs in about 10% of all test results. Methods: A large pedigree with 1 affected patient carrying a 71 and a 51 CA G repeat and 2 asymptomatic relatives carrying the 51 CAG repeat and normal -size alleles underwent clinical and molecular studies. Intragenic haplotyp es for these alleles were determined, A representative sample of the health y population in the region was obtained to assess the distribution of the n ormal (CAG), length. We established the genotype for 4 intragenic polymorph isms in the gene for MJD (MJD1) in 21 homoallelic individuals, to distingui sh their 2 normal chromosomes. In addition, we developed a new Southern blo t method to completely exclude cases of nonamplification of expanded allele s in the homoallelic individuals. Results: The study of the family in which the 51 CAG repeat was found sugge sts that the allele is apparently associated with disease. These intermedia te alleles were not present in a large sample of the healthy from the same region. Intragenic polymorphisms allowed distinction of the 2 different nor mal alleles in cases of homoallelism, The absence of an expanded allele was also confirmed by Southern blot. Conclusions: We propose an improved protocol for molecular testing for MJD. These strategies, developed to overcome the practical difficulties mostly in the presymptomatic and prenatal diagnosis of MJD, should useful for othe r polyglutamine -related disorders.