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目的:分析我国脊肌萎缩症(SMA)患儿SMN2基因甲基化水平与其转录水平,并初步探讨该基因的甲基化修饰是否影响我国儿童型SMA疾病的严重程度。创新点:首次在中国儿童型SMA人群中分析SMN2基因甲基化状态,提示该基因的甲基化模式在不同人种间具有一定保守性,而甲基化单元的甲基化状态可能具有种族差异。本研究也初步提示SMN2基因甲基化水平除了可能影响该基因的转录,还可能影响基因的可变剪接。方法:应用MassA RRAY的方法检测35例SMA患儿(SMN1基因纯合缺失,SMN2基因3拷贝)外周血细胞中SMN2基因甲基化状态;应用实时聚合酶链反应(real-time PCR)的方法检测SMN2基因不同转录本的表达水平;分析SMN2基因甲基化与该基因的转录以及SMA疾病严重程度的关系。结论:位于甲基化岛1的两个甲基化单元(nt-871和nt-735)和位于甲基化岛4的nt+999甲基化单元的甲基化水平在III型患儿中显著低于II型和I型患儿;nt-871和nt-290/-288/-285甲基化单元的甲基化水平与SMN2基因全长转录本(SMN2-fl)的转录水平呈负相关。此外,nt+938甲基化单元的甲基化水平与SMN2基因全长转录本与跳跃外显子7转录本的比值(fl/?7)呈负相关,但与SMN2的转录水平无关。因此,我们初步得出SMN2基因甲基化可能通过调控其转录而影响我国儿童型SMA的疾病表型。
Objective: To analyze the methylation level and transcription level of SMN2 gene in children with spinal muscular atrophy (SMA) in our country and to investigate whether the methylation of this gene affects the severity of childhood SMA in our country. Innovative point: The first analysis of the methylation status of SMN2 gene in Chinese children with SMA showed that the methylation pattern of this gene is somewhat conservative among different populations, and the methylation status of methylation unit may have a race difference. This study also preliminary suggests that SMN2 gene methylation level may affect the alternative splicing of genes in addition to possibly affecting the transcription of the gene. Methods: The methylation status of SMN2 gene in peripheral blood cells of 35 cases of SMA (homozygous deletion of SMN1 gene and 3 copies of SMN2 gene) was detected by MassA RRAY method. Real-time PCR was used to detect the methylation status of SMN2 gene. SMN2 gene expression in different transcripts; analysis of SMN2 gene methylation and the gene transcription and the severity of SMA disease. Conclusions: The methylation levels of two methylated units (nt-871 and nt-735) located on methylated island 1 and nt + 999 methylated units located on methylated island 4 are present in type III children Was significantly lower than that of type II and type I children. Methylation levels of nt-871 and nt-290 / -288 / -285 methylation units were negatively correlated with SMN2-SMN2-fl transcripts Related. In addition, the methylation level of nt + 938 methylation units was negatively correlated with the ratio of SMN2 full-length transcripts to hop-7 transcripts (fl /? 7), but not with SMN2 transcript levels. Therefore, we initially conclude that methylation of SMN2 gene may affect the phenotype of childhood SMA in our country by regulating its transcription.