探讨江苏省苏州市苯丙酮尿症(PKU)患儿苯丙氨酸羟化酶(PAH)基因突变特点。

选择2008－2011年，于上海新华医院通过串联质朴技术、苯丙氨酸负荷试验及尿蝶呤检测确诊分型的江苏省苏州市内25例PKU患儿及其父母为研究对象，通过SNaPShot基因分型技术和二代测序(NGS)技术，检测25例PKU患儿及其父母的PAH基因的突变。采用Sanger测序验证上述2种测序的结果，并分析苏州市PKU患儿PAH基因突变特点。本研究遵循的程序符合南京医科大学附属苏州医院人体试验委员会所制定的伦理学标准，得到该委员会批准，并与患儿监护人签署临床研究知情同意书。

SNaPShot基因分型技术、NGS技术检测本组受试者PAH基因突变结果与Sanger测序结果一致。25例PKU患儿中，运用SNaPShot基因分型技术，共检测出PAH基因的9种突变类型，分别为Exon7-R243Q、Exon6-Y204X、Exon7-R241C、Exon5-IVS4-1、Exon12-R413P、Exon3-R111X、Exon7-R261Q、Exon10-W326X及Exon11-Y356X，PAH基因突变检出率为58.0%(29/50)。其中10例患儿可通过SNaPShot基因分型技术检测到PAH基因纯合突变或复合杂合突变，家系分析可提供100%的基因诊断信息。对另15例不能提供100%基因诊断信息的PKU患儿运用NGS技术进行检测，在其中12例患儿中检测到PAH基因复合杂合突变，家系分析可提供100%的基因诊断信息；其余3例患儿仅检测到PAH基因单位点杂合突变。运用NGS技术发现了PAH基因的另外15种突变，包括突变频率为4%的A434D、V399V和Y325X 。上述2种方法对PAH基因总突变的检出率为94%(47/50)。

江苏省苏州市PKU患儿的PAH基因热点突变包括R243Q、Y204X、R241C、IVS4-1、R413P、R111X、A434D、V399V和Y325X。运用SNaPShot基因分型技术检测PAH基因突变，具有一定临床应用价值。

To study the characteristics of phenylalanine hydroxylase(PAH) gene mutations in children with phenylketonuria(PKU) in Suzhou, Jiangsu Province.

From 2008 to 2011, a total of 25 cases PKU children in Suzhou, Jiangsu Province, who were confirmed genotyping by tandem mass spectrometry, phenylalanine loading test and uropterin test with their blood sample in Shanghai Xinhua Hospital, and their parents were all enrolled as the research objects.Mutations in PAH gene of 25 children with PKU and their parents were detected by SNaPShot genotyping technology and next generation sequencing(NGS) technology.The results of genotypes by these two technology were verified by Sanger sequencing.The characteristics of PAH gene mutations of PAH gene in children with PKU in Suzhou, Jiangsu Province were analyzed. The study protocol was approved by the Ethical Review Board of Investigation in Human Being of Nanjing Medical University Affiliated Suzhou Municipal Hospital. Informed consent was obtained from the parents of each participating child.

The results of PAH gene mutations in the subjects by SNaPShot genotyping and NGS technology were coincide with the Sanger squencing results.Using SNaPShot genotyping technology, 9 kinds of mutations of PAH gene in 25 children with PKU could be detected, which including Exon7-R243Q, Exon6-Y204X, Exon7-R241C, Exon5-IVS4-1, Exon12-R413P, Exon3-R111X, Exon7-R261Q, Exon10-W326X and Exon11-Y356X, and PAH gene mutations detection rate was 58.0%(29/50). Ten of 25 children with PKU carrying PAH gene homozygous mutations or compound heterozygous mutations could be detected by SNaPShot genotyping technology, and pedigree analysis can confirm 100% genetic diagnosis information.Another 15 of 25 children with PKU tested by SNaPShot genotyping technology carrying one PAH gene heterozygous mutation or not, were further tested by NGS technology, and 12 of 15 children could be detected carrying PAH gene compound heterozygous mutations, but another 3 of 15 children, only could be detected carrying PAH gene heterozygous mutation.Other 15 kinds of mutations were detected by NGS technology, including A434D、V399V and Y325X, with 4.0% mutation frequency.The PAH gene total mutation detection rate of combined SNaPShot genotyping and NGS technology was 94.0%(47/50).

The hot spot mutations in PAH gene of children with PKU in Suzhou, Jiangsu Province included R243Q, Y204X, R241C, IVS4-1, R413P, R111X, A434D, V399V and Y325X.Testing PAH gene mutations by SNaPShot genotyping technology is a helpful method to gene diagnosis of PKU.