Weijiang Jin1, Zhengquan Yang1, Xiaojun Tang2, Xiuchao Wang1, Yaxin Huang1, Chenmin Hui1, Jiaming Yao1, Ju Luan3, Shaohua Tang4, Shengnan Wu5, Shengnan Jin6, Chunming Ding7. 1. School of Laboratory Medicine and Life Science, Wenzhou Medical University, Wenzhou, Zhejiang, China; Key Laboratory of Laboratory Medicine, Ministry of Education, Wenzhou Medical University, Wenzhou, Zhejiang, China. 2. Molecular Diagnostic Laboratory, Shanghai Children's Hospital, Shanghai Jiaotong University, Shanghai, China. 3. Key Laboratory of Laboratory Medicine, Ministry of Education, Wenzhou Medical University, Wenzhou, Zhejiang, China; InnoMed Diagnostics Inc, Wenzhou, China. 4. Key Laboratory of Birth Defects, Department of Genetics, Wenzhou Central Hospital, Wenzhou, China. 5. Molecular Diagnostic Laboratory, Shanghai Children's Hospital, Shanghai Jiaotong University, Shanghai, China. Electronic address: wushengnan@shchildren.com.cn. 6. School of Laboratory Medicine and Life Science, Wenzhou Medical University, Wenzhou, Zhejiang, China; Key Laboratory of Laboratory Medicine, Ministry of Education, Wenzhou Medical University, Wenzhou, Zhejiang, China. Electronic address: snjin1997@qq.com. 7. School of Laboratory Medicine and Life Science, Wenzhou Medical University, Wenzhou, Zhejiang, China; Key Laboratory of Laboratory Medicine, Ministry of Education, Wenzhou Medical University, Wenzhou, Zhejiang, China. Electronic address: cmding@gmail.com.
Abstract
BACKGROUND: Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder caused by defects in the survival motor neuron 1 (SMN1) gene. Homozygous deletion of the SMN1 gene accounts for 95% of all affected SMA patients. A highly homologous gene survival motor neuron 2 (SMN2) compensates weakly with the loss of SMN1 and its copy number correlates with disease severity. METHODS: We report here the MS-CNV method combining competitive PCR and MALDI-TOF mass spectrometry for simultaneous quantification of SMN1, SMN2 and NAIP dosages. For both SMN1 and SMN2, the exon 7 and exon 8 were analyzed. MS-CNV was validated with parallel analysis by a commercial MLPA assay in two independent cohorts. RESULTS: In the first cohort of 79 blood samples containing 3 SMA patients and 5 carriers, MS-CNV results were highly concordant with MLPA analysis for the copy numbers of SMN1, SMN2 and NAIP. In the second independent and blinded cohort of 62 blood samples containing 21 SMA patients and 14 carriers, MS-CNV results were also highly concordant with MLPA. Both MS-CNV and MLPA quantified SMN1 dosages without ambiguity. CONCLUSIONS: MS-CNV can be used for carrier screening and genetic diagnosis of SMA, providing dosages information for both SMN1 and SMN2 given its accuracy and high sample processing throughput by mass spectrometric analysis.
BACKGROUND: Spinal muscular atrophy (SMA) is an autosomal recessive neurodegenerative disorder caused by defects in the survival motor neuron 1 (SMN1) gene. Homozygous deletion of the SMN1 gene accounts for 95% of all affected SMA patients. A highly homologous gene survival motor neuron 2 (SMN2) compensates weakly with the loss of SMN1 and its copy number correlates with disease severity. METHODS: We report here the MS-CNV method combining competitive PCR and MALDI-TOF mass spectrometry for simultaneous quantification of SMN1, SMN2 and NAIP dosages. For both SMN1 and SMN2, the exon 7 and exon 8 were analyzed. MS-CNV was validated with parallel analysis by a commercial MLPA assay in two independent cohorts. RESULTS: In the first cohort of 79 blood samples containing 3 SMA patients and 5 carriers, MS-CNV results were highly concordant with MLPA analysis for the copy numbers of SMN1, SMN2 and NAIP. In the second independent and blinded cohort of 62 blood samples containing 21 SMA patients and 14 carriers, MS-CNV results were also highly concordant with MLPA. Both MS-CNV and MLPA quantified SMN1 dosages without ambiguity. CONCLUSIONS: MS-CNV can be used for carrier screening and genetic diagnosis of SMA, providing dosages information for both SMN1 and SMN2 given its accuracy and high sample processing throughput by mass spectrometric analysis.