Background: Multiplex ligation probe amplification (MLPA) is a sensitive and efficient technique for molecular diagnosis of diseases involving deletions or duplications of large genomic regions. In phenylketonuria (PKU), most of the mutant alleles correspond to missense mutations and large deletions have been scarcely identified. In this study, we report for the first time the use of MLPA analysis on PKU patients to detect exonic deletions.
Method: DNA from 22 unrelated PKU patients with an incomplete genetic diagnosis after standard mutation detection analysis were subjected to MLPA analysis. Deletions were confirmed by long-range PCR and sequence analysis.
Results: The technique identified two large genomic deletions in the phenylalanine hydroxylase (PAH) gene, of 6.6 kb and 1.8 kb, including exons 3 and 5, respectively. The chromosomal breakpoints were established by long-range PCR and chromosomal walking, confirming the involvement of repetitive sequences in the deletions.
Conclusion: MLPA may complement routine mutation screening in PKU patients, although, in the sample studied, exonic deletions in the PAH gene do not appear to be a frequent cause of PKU.