MLPA方法在脊髓性肌肉萎缩症分子诊断中的应用

目的 探讨多重连接依赖性探针扩增(MLPA)技术在脊髓性肌肉萎缩症(SMA)分子诊断中的应用.方法 从13例SMA患者、31名患者父母的外周血标本和10份胎儿羊水标本,以及50名正常人外周血标本中提取基因组DNA,应用MLPA技术进行分析,同时也行常规聚合酶链反应-限制性片段长度多态性(PCR-RFLP)和位点特异性PCR分析.结果 MLPA分析结果与常规PCR-RFLP和位点特异性PCR结果相符:13例患者的运动神经元存活基因(SMN)1基因均呈纯合缺失,SMN2基因拷贝数的增加与SMA表型的严重程度(从I型到Ⅲ型)存在显著性差异(P<0.05);31名患者父母SMN1基因1拷贝的人数为29(占93.5%),2拷贝的为2(占6.5%);50名正常健康成人SMN1基因1拷贝的人数为1(占2.0%),2拷贝的为48(占96.O%);SMA患者父母组和健康正常成人组之间的SMN1基因拷贝数存在显著件差异(P<0.01);10例胎儿中2例存在SMN1的纯合缺失.结论 MLPA是一种准确可靠的SMA分子诊断新方法.

Molecular diagnosis of spinal muscular atrophy by multiplex ligation-dependent probe amplification

Objective To investigate the effect of multiplex ligation-dependent probe amplification (MLPA)in molecular diagnosis of spinal muscular atrophy(SMA).Methods Peripheral blood samples were collected from 13 SMA patients.31 parents of SMA patients,50 healthy individuals without family history of SMA,and 10 specimens of amniotic fluid from these families were collected too.Genomic DNA was analyzed by MLPA,conventional PCR-RFLP,and allele-specific PCR.Results In complete agreement with the results of conventional PCR-RFLP and allele-specific PCR.MLPA analysis showed that all of the 13 patients had homozygous deletion of the Survival of motor neuron 1(SMN1)geBe,and there Wag significant difference between the SMA severity(type I to typeⅢ)and SMN2 copy humber(P<0.05).of the 31 parents 29(93.5%)had 1 copy of SMNI,2(6.5%)had 2 copies of SMN1.Of the 50 healthy individuals.1(2.0%)had 1 copy of SMN1,48(96.O%)had 2 copies of SMN1,and 1(2.0%)had 3 copies.The SMN1 copy humber of the parents was significantly higher than that of the healthy individuals (P<0.01).Two of the 10 feruses had homozygous deletion of SMN1.Conclusion The MLPA technique has proved to be an accurate and reliable tool for the molecular diagnosis of SMA.both in patients and in healthy carriers.