Abstract
ObjectiveArgininosuccinate lyase (ASL) gene mutations account for argininosuccinic aciduria (ASA). This study aimed to design a minigene construct of ASL gene in order to investigate the impact of variants on splicing.MethodsThe peripheral blood samples were collected from the family members, and genomic DNA was extracted for gene diagnosis using the total exon sequencing method. The novel mutation gene was cloned into pEGFP-C1 vector, and the pathogenicity of the mutation was examined in cultured cells in vitro.ResultsThe clinical diagnosis of the proband as ASA was clear. Two pathogenic mutations, c.281G>T (p.Arg94Leu) and c.208-15 T>A were detected in the ASL gene, and the two mutations had not been reported. The minigene expression in vitro confirmed that c.208-15 T>A could cause aberrant splicing, resulting in the retention of 13 bp in intron 2 to exon 3.ConclusionTwo new pathogenic mutations of ASL gene, c.208-15 T>A and c.281G>T, were found in an ASA family, which enriches the mutational profile of the ASL gene and provides a basis for genetic diagnosis of ASA. Minigenes are optimal approaches to determine whether the intron mutation can cause aberrant splicing.
Highlights
Argininosuccinic aciduria (ASA) is a clinically rare urea cycle disorder, and belongs to the autosomal recessive genetic disease
The variant in intron can be examined by mini-gene splicing analysis (Bonnet et al, 2008; Théry et al, 2011)
We report two novel mutations in argininosuccinate lyase (ASL) gene in ASA patient, one of which is on the intron, and we functionally characterized the mutations
Summary
Argininosuccinic aciduria (ASA) is a clinically rare urea cycle disorder, and belongs to the autosomal recessive genetic disease. ASA was first reported by ALLAN in 1958 (Allan et al, 1958). The overall reported prevalence of ASA is 1:70,000 (Nagamani et al, 2012c). ASA is due to the defect of argininosuccinate lyase (ASL) gene which causes arginyl succinic acid not to be cleaved into arginine and fumarate. ASA can be divided into neonatal type and delayed type according to the onset of disease. The neonatal ASA is severe and the mortality rate is high (Chen et al, 2010). The first case of neonatal ASA in China was reported in 2014, and the diagnosis was confirmed by genetic testing after death (Yuan et al, 2014).
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