Rapid touchdown PCR assay for the molecular diagnosis of spinocerebellar ataxia type 2

Seven different chromosomal loci, designated SCAI to SCA7 (spinocerebellar ataxias), have been identified as responsible for autosomal dominant cerebellar ataxias. Five genes (SCA1, 2, 3, 6, 7) have been cloned to date and show a single type of mutation, an unstable expansion of a CAG repeat coding for a polyglutamine stretch in the corresponding protein. We describe an improved polymerase chain reaction assay, based on a touchdown protocol, for the diagnosis of spinocerebellar ataxia type 2. This method produces an efficient amplification of both normal and pathological alleles and no radioactive labelling is necessary to observe the amplification products. The pathological alleles are identified by a simple non-denaturing polyacrylamide electrophoretic separation followed by ethidium bromide staining. A comparison of this technique with previously reported methods confirmed its utility for the rapid molecular diagnosis of spinocerebellar ataxia type 2. We found that the spinocerebellar ataxia type 2 mutation is responsible for 88% of the examined autosomal dominant cerebellar ataxia type 1 families in our territory (eastern Sicily). With the rapid touchdown polymerase chain reaction method, the trinucleotide expansion was also observed in 2 ataxic patients without family history of the disease, suggesting the necessity for analysis of spinocerebellar ataxia type 2 expansion even in sporadic patients.