数字PCR技术检测脊髓性肌萎缩症携带者的应用评估  

An evaluation of carrier detection for Spinal muscular atrophy using digital PCR assay

作　　者：闫有圣 陈芊如 张萌[1] 王芳 王一鹏[1] 陈昕雯 阴赪宏[1] 郭永[2] Yan Yousheng;Tan Chianru;Zhang Meng;Wang Fang;Wang Yipeng;Chen Xinwen;Yin Chenghong;Guo Yong(Prenatal Diagnostic Center,Beijing Obstetrics and Gynecology Hospital,Capital Medical University,Beijing Maternal and Child Health Care Hospital,Beijing 100026,China;Department of Biomedical Engineering,School of Medicine,Tsinghua University,Beijing 100084,China)

机构地区：[1]首都医科大学附属北京妇产医院/北京妇幼保健院,北京100026 [2]清华大学医学院生物医学工程系,北京100084

出　　处：《中华医学遗传学杂志》2024年第1期20-24,共5页Chinese Journal of Medical Genetics

基　　金：首都临床特色诊疗技术研究及转化应用项目(Z221100007422012);国家自然科学基金(81902157)。

摘　　要：目的应用数字PCR方法进行脊髓性肌萎缩症(SMA)携带者检测,评价其效果以及将其用于SMA携带者筛查的可行性。方法收集常规进行产前SMA携带者筛查的孕妇外周血样本214例,其中204例为随机样本,10例为已知SMN1第7和第8外显子拷贝数的样本,随机掺入实验中,用于对数字PCR方法进行性能验证。采用数字PCR方法对外周血DNA样本中SMN1和SMN2基因的第7和第8外显子拷贝数进行检测,并与荧光定量PCR的检测结果进行比较,评估数字PCR方法的可靠性和检测效能。结果采用数字PCR方法在204例随机样本中检测到SMN1第7和第8外显子同时杂合缺失样本5例,仅SMN1第8外显子杂合缺失样本3例,SMN1第7和第8外显子未缺失样本196例。10例已知SMN1第7和第8外显子拷贝数样本的检测结果与预期值一致。通过与荧光定量PCR检测结果比较,数字PCR对于SMN1第7和第8外显子拷贝数的检测结果的一致率为100%。结论数字PCR检测SMN1第7和第8外显子的缺失情况结果与qPCR一致,并且能够明确区分目标基因的拷贝数,该方法可用于SMA致病基因的携带者筛查。此外,数字PCR还能检测SMN2第7和第8外显子的拷贝数,提供更多的信息。Objective To assess the effectiveness and feasibility of carrier detection for Spinal muscular atrophy(SMA)by using digital PCR assay.Methods Peripheral blood samples were collected from 214 pregnant women who were routinely screened for SMA carriers,of which 204 were randomly selected samples and 10 were samples with known copy numbers of SMN1 exons 7 and 8.Samples with known copy numbers of SMN1 exons 7 and 8 were randomly mixed into the experiment to validate the performance of the digital PCR assay.The copy numbers of SMN1 exons 7 and 8 and SMN2 exons 7 and 8 in peripheral blood samples were detected by digital PCR assay.The results of SMN1 exons 7 and 8 were compared with those of the quantitative PCR method to assess the reliability and clinical performance of the digital PCR assay.Results Among the 204 random samples,digital PCR has detected five samples with simultaneous heterozygous deletion of SMN1 exons 7 and 8,three samples with heterozygous deletion of SMN1 exon 8 only,and 196 samples with no deletion of SMN1 exons 7 and 8.Ten samples with known SMN1 exons 7 and 8 copy numbers were detected with the expected values.The digital PCR test results were fully consistent with that of the quantitative PCR.Conclusion The results of digital PCR for the detection of copy number variation of SMN1 exons 7 and 8 were consistent with qPCR.Digital PCR assay was able to clearly distinguish the copy number of the target genes,therefore can be used for SMA carrier screening.Moreover,it can also detect copy number of SMN2 exons 7 and 8,which can provide more information for genetic counseling.

关 键 词：脊髓性肌萎缩症 荧光定量PCR 数字PCR 携带者筛查 

分 类 号：R714.5[医药卫生—妇产科学]