北京地区苯丙酮尿症基因突变构成及基因型与表型相关分析

目的 明确北京地区苯丙酮尿症(phenylketonuria,PKU)患儿苯丙氨酸羟化酶基因(phylalanine hydroxylase gene,PAH)突变图谱及其突变基因的微单体型(STR/VNTR)构成,探讨突变基因型与生化代谢表型的相关关系.方法 应用PCR/SSCP、序列分析和变性凝胶电泳等技术,对50例北京地区PKU患儿及其父母,进行PAH基因的全部外显子及两侧的内含子序列和STR与VNTR多态性分布的分析.依据由基因型预测生化代谢表型的方法进行基因型与生化代谢表型的相关关系分析.结果 (1)共检测到34种PAH突变基因,总的检出率为95%;较为常见的突变有R243Q(20%)、EX6-96A＞G(11%)、Y356X(9%)和V399V(7%),其次是R111x(5%)、R413P(5%)、R252Q(3%)和A434D(3%).(2)北京地区PKU患儿的STR杂合度较高,共检测出8种等位基因,以240 bp(34%)和244 bp(44%)最为常见;而VNTR的杂合度较低,只检测到3种等位基因,以VNTR3(83%)最为常见.(3)由基因型预测的生化代谢表型与患儿实际的生化代谢表型之间一致率为81.5%,在经典型PKU中一致率达到87.5%.结论 (1)北京地区共有34种PAH基因突变,检出率为95%,以R243Q、EX6-69A＞G、Y356X和V399V突变为常见突变.(2)PAH突变基因的微单位(STR/VNTR)构成以240/3和244/3最为常见.(3)PKU患儿基因型与生化代谢表型之间存在较好的相关关系,一致率达到80%以上.

Spectrum of phylalanine hydroxylase gene mutations and genotype-phenotype correlation in the patients with phenylketonuria in Beijing area of China

Objective To identify the mutation spectrum and the distribution of minihaplotypes (STR/VNTR)of phenylalanine hydroxylase(PAH)gene and explore the C01Te]ations between genotype and phenotype of patients with phenylketonuria(PKU)in Beijing area of China.Method (1)Fifty cases with PKU were involved in this study.PKU was identified by the Neonatal Screening Center of Beijing.All 13 exons and their flanking intronic sequences of PAH gene of these patients were amplified and then subjected to SSCP analysis and direct sequencing.(2)The distribution of polymorphic locus of short tandem repeat (STR)and variable number tandem repeat(VNTR)was analyzed by PCR and denaturing gel electrophoresis.(3)The correlations between genotype and phenotype were studied by analysis of the matching rate between the expected and observed phenotypes.The predicted phenotype was determined on the basis of the sum of the assigned values of the two mutant alleles.Results (1)A total of 34 different mutations were detected with the relative frequency of 95% among 50 PKU patients.The prevalent mutations in this study were:R243Q(20%),EX6-96A＞G(11%),Y356X(9%),and V399V(7%).The next common mutations were R111X(5%),R413P(5%),R252Q(3%)and A434D(3%).Thirty-four detected mutations were distributed throughout the whole PAH gene,except exon 1,8 and 13.Exon 7 and 11,with the mutant rate 34% and 19% respectively,seemed to be the hot mutant areas/regions of PAH gene.(2)The minihaplotypes(STR/VNTR)of 34 mutations were identified in this research.The STR and VNTR showed 8 and 3 alleles,respectively.Among them,244 bp(44%)and 240 bp(34%)were the prevalent STR alleles.Meanwhile,the VNTR3(83%)was the most common VNTR allele in PKU patients.(3)A better consistency(81.5%)between expected and observed phenotypes was revealed by analysis of correlation between genotype and phenotype.Especially in classic PKU,the consistency rate was up to 87.5%.Conclusion(1)The frequency distribution of common PAH gene mutations in Beijing region was close to that of Tianjin and Yunnan regions,while it was different from that of Southern regions of China,such as Guangzhou.especially Taiwan.The PAH mutation with a highly heterogeneous trait was also demonstrated in this study.(2)STR and VNTR minihaplotype will prove helpful to trace the origins of PAH mutations and to analyze the genetic drift.However,the most minihaplotypes of the STR/VNTR are similar,so it is necessary to associate some other polymorphic loci with the STR/VNTR minihaplotype to analyze the different mutations.(3)The fact that a better consistency existed between phenotypes and genotype with most PKU patients suggested that the study of the genotype of PKU patients would be helpful to the individualized treatment and to genetic counseling for their families.