1. Introduction
Phenylketonuria (PKU) is an autosomal recessive metabolic disease caused by mutations in the gene encoding phenylalanine hydroxylase (PAH) (1). At present, the early diagnosis and intervention of PKU through neonatal disease screening can avoid the damage to the nervous system and other tissues caused by PAH mutations (2). At the same time, prenatal diagnosis can reduce the chance of giving birth to children with PKU in high-risk families. From the perspective of the three-level prevention system, the most effective way to prevent PKU is to identify carriers before pregnancy, so as to take effective measures to avoid the birth of PKU children (3). However, due to the variety and heterogeneity of PAH gene mutations, a lot of basic and clinical research is needed to avoid the birth of children with PKU.
As the incidence of PKU and the spectrum of PAH gene mutations in Shaanxi province, China are not clear, it is necessary to combine PKU prevention strategies with the screening and diagnosis of PAH gene mutations, so as to form an effective three-level prevention model and improve the efficiency of PKU prevention. For gene diagnosis of PKU, the most commonly used detection technologies at present mainly include DNA sequencing, multiplex ligation-dependent probe amplification (MLPA), short tandem repeat linkage analysis, etc. However, there are problems associated with the use of these approaches, such as complex operation, high cost, high requirements for laboratory personnel, and not being conducive to popularization (4-5). Therefore, the purpose of this study was to develop amplification refractory mutation system-polymerase chain reaction (ARMS-PCR) combined with fluorescent probe technology to detect the mutation hotspots of the PAH gene, in order to develop a PAH gene screening kit that is convenient, fast, cheap, accurate, and suitable for all levels of screening agencies, thereby providing technical support for the “one-level prevention” of PKU (3).