基因诊断红细胞丙酮酸激酶缺乏症及PKLR基因新突变

Genetic diagnosis of erythrocyte pyruvate kinase deficiency and discovery of PKLR gene novel mutations

目的 通过对3例红细胞丙酮酸激酶缺乏症（pyruvate kinase deficiency,PKD）患儿临床症状及PKLR基因新突变类型分析,探讨PKD的基因诊断方法.方法 应用目标序列捕获和高通量测序技术对3例PKD患儿的血液系统疾病相关基因进行测序,并预测基因突变对蛋白质功能的影响,确定患儿的致病基因型后,应用Sanger测序技术对此基因型进行验证.结果 在3例患儿中发现了5种PKLR基因突变类型,分别为c.1529G ＞A（p.RS10Q）和c.1031T＞ G（p.I344S）复合杂合突变、c.847G＞ T（p.V283F）纯合突变,c.979delC（p.L327fs）和c.604_617del（p.V202fs）复合杂合突变,其中c.1529G＞ A（p.R510Q）突变已有文献报道,其余4种突变为PKLR基因新的突变类型.c.1031T＞G（p.I344S）、c.847G＞T（p.V283F）为可能致病变异,该基因突变致病的可能性＞90％;c.979delC （p.L327fs）和c.604_617del（p.V202fs）变异为致病变异.且这5种突变经预测对蛋白质功能影响较大,均为致病突变.结论 对于临床诊断困难的PKD患儿,可通过目标序列捕获和高通量测序技术发现PKLR基因变异,并评价其致病性.

Objective To investigate the clinical symptoms of pyruvate kinase deficiency （PKD） and the new mutation type of PKLR gene in 3 cases of PKD,and to explore the method for PKD gene diagnosis.Methods Sequencing of blood system-related genes in 3 children was performed by target sequence capture and high-throughput sequencing technology,and the protein function of mutant gene was forecasted,after detecting the pathogenicity of the patients,these genotypes were confirmed by Sanger sequencing.Results In the 3 children,5 types of PKLR gene mutations were found：double heterozygous mutations c.1529G 〉 A（p.R510Q） and c.1031T 〉 G（p.I344S）,homozygous mutation c.847G 〉 T （p.V283F）,double heterozygous mutations c.979delC （p.L327fs）and c.604_617del （p.V202fs）.PKLR gene c.1529G 〉 A（p.R510Q） mutation had been reported previously,and the other four mutations were new.c.1031T 〉 G （p.I344S） and c.847G 〉 T （p.V283F） was possibly pathogenic mutation,which meant that the probability of mutation of this gene was more than 90% and c.979delC （p.L327fs） and c.604 _617del （p.V202fs） variation was a pathogenic variation.These 5 mutations had a greater effect on protein function,and all ofthem were pathogenic mutations.Conclusion Since PKD patients are difficult to be diagnosed clinically,PKLR gene variation can be detected by target sequence capture and high throughput sequencing technology,and the pathogenicity of the new mutant is evaluated.