DHPLC技术在儿童型脊髓性肌萎缩症的基因诊断及携带者基因筛查中的应用

The Application of DHPLC in the Gene Diagnosis of the Childhood Type Spinal Muscular Atrophy and in the Gene Screening of the SMA Carriers

目的评价变性高效液相色谱(denaturing high performance liquid chromatography,DHPLC)技术在儿童型脊髓性肌萎缩症(spinal muscul aratrophy,SMA)基因诊断及携带者基因筛查中的应用价值。方法对35例临床疑诊为儿童型SMA的患者采用临床标准进行诊断。同时,应用DHPLC技术联合双重PCR对所有入选患者、临床标准确诊为SMA患者的双亲以及一份标准对照进行SMN1拷贝数检测,做出基因诊断及判断携带者。结果(1)35例入选患者,临床标准诊断确诊SMA15例,非SMA20例。(2)DHPLC技术检测35例入选患者:15例临床诊断SMA患者中,12例SMN1拷贝数为0,为纯合缺失型SMA患者;2例SMN1拷贝数为1,为杂合缺失型SMA患者;1例SMN1拷贝数为2,为非SMA。20例临床诊断非SMA者,SMN1拷贝数为2,为非SMA。(3)与临床标准诊断相比,DHPLC技术诊断SMA灵敏度为93.3%,特异度为100%。15例临床确诊为SMA患者的28名双亲中,1例SMN1拷贝数为0,6例SMN1拷贝数为2,21例SMN1基因拷贝数为1(为携带者),SMA患者双亲的携带率为75%。结论应用DHPLC技术可以检测出SMN1基因拷贝数,进行SMA基因诊断灵敏度和特异度高,适合临床应用;应用该技术可以进行携带者筛查,为遗传分析和产前诊断提供理论依据。

Objective To assess the practical value of denaturing high performance liquid chromatography （DHPLC） in gene diagnosis of the childhood type spinal muscular atrophy （SMA） and gene screening of the SMA carriers. Methods The 35 selected children suspicious of childhood type SMA were diagnosed with clinical standard. The SMN1 copy numbers of the selected children and their parents and one standard sample were detected by DHPLC combined with double-PCR. Results （1） Diagnosed by clinical standard, 15 children of the 35 selected children were SMA patients and 20 children were non-SMA patients. （2） Detected by DHPLC,the results of the 35 selected children＇s samples were as follows： of the 15 clinically diagnosed SMA children, 12 were homozygous deletion type SMA patients carrying 0 copy of SMN1 ;2 were heterozy-gous deletion type SMA patients carrying 1 copy of SMN1;1 was non-SMA patient having 2 cop- ies of SMN1. The 20 clinically diagnosed non-SMA children had 2 copies of SMN1. （3）Compared with the clinical standard,the sensitivity of the DHPLC is 93.3% ,and the specificity is 100%. （4） Of the parents＇ samples of the 15 clinical diagnosed SMA patients, 1 had 0 copy,6 had 2 copies,21 had 1 copy of SMNI（SMA carriers）. The carrier incidence of the SMA children＇s parents was 75 ~. Conclusion It suggests that DHPLC could detect SMN1 copy numbers with high sensitivity and specificity, thus quite suitable for practical application of gene diagnosis of SMA, so that it could be applied to screening of SMA carriers, which could in turn provide essential data for genetic counseling and prenatal diagnosis.