CAJ | 학술논문

目的探讨聚合酶链反应-限制性片段长度多态性(polymerase chain reaction-restriction fragment length polymorphism,PCR-RFLP)技术在脊髓性肌萎缩症(spinal muscular atrophy,SMA)基因诊断中的应用.方法用PCR-RFLP分析935例临床疑似SMA患儿的运动神经元存活基因1(survival motor neuron,SMN1)第7和第8外显子的缺失,同时用多重连接探针扩增技术(multiplex ligationdependent probe amplification,MLPA)分析其中339例疑似病例的SMN1基因拷贝数改变.用Pearson卡方检验分析两种方法检测SMN1纯合和杂合性缺失的一致性.结果共发现SMN1基因第7外显子纯合缺失590例,疑似患者的SMA基因诊断率为63.1％(590/935).用PCR-RFLP和MLPA技术联合分析疑似病例339例,PCR-RFLP共发现SMN1纯合缺失194例,MLPA发现196例,二者的一致性为98.9％,差异无统计学意义(x2=0.2,P=0.88).PCR-RFLP仅发现SMN1疑似杂合性缺失4例,而MLPA证实有17例,二者的一致性为23.5％,差异有统计学意义(x2=8.29,P＜0.01).结论 PCR-RFLP尽管简便、特异、实用,但对于5％～10％ SMN1杂合缺失合并点突变的病例则存在明显的局限性.
목적 탐토취합매련반응-한제성편단장도다태성(polymerase chain reaction-restriction fragment length polymorphism,PCR-RFLP)기술재척수성기위축증(spinal muscular atrophy,SMA)기인진단중적응용.방법 용PCR-RFLP분석935례림상의사SMA환인적운동신경원존활기인1(survival motor neuron,SMN1)제7화제8외현자적결실,동시용다중련접탐침확증기술(multiplex ligationdependent probe amplification,MLPA)분석기중339례의사병례적SMN1기인고패수개변.용Pearson잡방검험분석량충방법검측SMN1순합화잡합성결실적일치성.결과 공발현SMN1기인제7외현자순합결실590례,의사환자적SMA기인진단솔위63.1％(590/935).용PCR-RFLP화MLPA기술연합분석의사병례339례,PCR-RFLP공발현SMN1순합결실194례,MLPA발현196례,이자적일치성위98.9％,차이무통계학의의(x2=0.2,P=0.88).PCR-RFLP부발현SMN1의사잡합성결실4례,이MLPA증실유17례,이자적일치성위23.5％,차이유통계학의의(x2=8.29,P＜0.01).결론 PCR-RFLP진관간편、특이、실용,단대우5％～10％ SMN1잡합결실합병점돌변적병례칙존재명현적국한성.
Objective To explore the applicability and limitation of PCR-restriction fragment length polymorphism (PCR-RFLP) method for genetic diagnosis of spinal muscular atrophy (SMA).Methods PCR-RFLP was applied to detect potential deletion in exons 7 and 8 of SMN1 gene in 935 suspected cases with SMA.Multiplex ligation-dependent probe amplification(MLPA) was carried out to analyze dosage alteration of SMN1 gene in 339 of such cases. To confirm the accuracy of PCR-RFLP method for homozygous and heterozygous deletions detection,the consistency of PCR-RFLP and MLPA results were assessed with a Pearson Chi-square test.Results Homozygous deletion of exon 7 of SMN1 was detected in 590 suspected cases.The rate of diagnosis was therefore 63.1％ (590/935).For the 339 suspected cases,PCR-RFLP and MLPA respectively identified 194 and 196 homozygous deletions in the exon 7 of SMN1 gene,suggesting a good consistency (98.9％)(x2 =0.2,P=0.88).However,only 4 of 339 cases was found to carry a heterozygous deletion of SMN1 exon 7 by PCR-RFLP,in contrast with 17 detected by MLPA.The consistency only reached 23.5％,for which statistical significance was detected (x2 =8.29,P＜0.01).Conclusion Although PCR-RFLP is a simple,specific and efficient method for SMA diagnosis,it has obvious limitation for the diagnosis of 5％-10％ SMA patients who have carried a compound heterozygous mutation.