一例脊肌萎缩症患者及其家系的SMN基因突变分析

Mutation analysis of SMN gene in a patient and his family with spinal muscular atrophy

目的 对1例脊肌萎缩症（spinal muscular atrophy，SMA）患者及其家系成员的SMN基因行突变分析。方法 采用多重连接依赖性探针扩增（multiplex ligation-dependent probe amplification，MLPA）技术、逆转录聚合酶链反应及T克隆-测序技术对患者SMN基因进行拷贝数分析和点突变的鉴定，应用MLPA和针对点突变区域的SMN基因第5外显子PCR-直接测序法对患者父母SMN基因进行拷贝数分析和点突变的证实，同时用200名正常人外周血进行相关位点对照研究。结果 患者具有1个拷贝的SMN1基因和1个拷贝的SMN2基因，在这个SMN1基因第230位密码子上存在1个未见报道的错义突变S230L（TCA→TTA）；父亲具有两个SMN1和两个SMN2拷贝，其中1个SMN1基因存在S230L突变；母亲具有1个SMN1拷贝，而SMN2基因是纯合缺失的。200名对照中未发现该位点突变。结论 在1个SMA家系中发现了1个新的SMN1基因点突变，即S230L，并准确分析了此家系各成员的SMN基因型。

Objective To perform mutation analysis and describe the genotype of the SMN gene in a patient with spinal muscular atrophy （SMA） and his family. Methods Deletion analysis of the SMN1 exon 7 by conventional PCR-restriction fragment length polymorphism（RFLP） and allele-specific PCR, and gene dosage of SMN1 and SMN2 by multiplex ligation-dependent probe amplification （MLPA） were performed for the patient and his parents; reverse transcriptase （RT）-PCR and sequencing were performed for the patient. To determine whether the SMN variant was exclusive to transcripts derived from SMN1, the RTPCR product of the patient was subcloned and multiple clones were sequenced directly; PCR of SMN exon 5 from the genomic DNA of the parents and direct sequencing were performed to confirm the mutation. Results In SMN1 exon 7 deletion analysis, no homozygous deletion of the SMN1 was observed in the family; the gene dosage analysis by MLPA showed that the patient had 1 copy of SMN1 and 1 copy of SMN2, his father had 2 copies of SMN1 and 2 copies of SMN2, and his mother had 1 copy of SMN1 and no SMN2. A previously unreported missense mutation of S230L was identified from the patient and this mutation was also found in his father. Conclusion A novel missense mutation of S230L was identified in the SMA family and the genotype of the family members were in vestigated.