Multiplex ligation-dependent probe amplification (MLPA) detects large deletions in the MECP2 gene of Swedish Rett syndrome patients
Multiplex ligation-dependent probe amplification (MLPA) detects large deletions in the MECP2 gene of Swedish Rett syndrome patients
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DOI：
10.1089/109065703322783707
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发表时间：
2003-12-01
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影响因子：
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通讯作者：
Wahlström, J
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作者：
Erlandson, A;Samuelsson, L;Wahlström, J
Xq28上甲基CpG结合蛋白-2(MECP2)基因的突变被发现是Rett综合征(RS)的原因之一。在之前的突变筛查中，我们发现81%的瑞典经典Rett妇女存在MECP2突变。在这项研究中，我们使用多重连接依赖的探针扩增(MLPA)分析了22名患者的MECP2缺失。临床上11例为经典型的Rett女性，3例为先天性RS，7例为Rett变异型。作为纳入标准，我们使用了以前测序结果显示MECP2基因编码部分没有突变的患者的DNA。MLPA是一种基于多重聚合酶链式反应的方法。在一次聚合酶链式反应中，用相同的引物扩增了多达40个探针。扩增产物的特异性由扩增前每个探针结构的位点特异性杂交决定。每个PCR产物都有一个唯一的长度，这使得通过大小分离来识别它成为可能。在11例经典型Rett患者中，有3例(27%)使用MLPA检测到MECP2的大量缺失。所有患者均有两个外显子缺失，其中2例缺失涉及外显子3和4，1例缺失外显子1和2。在截形、先天性和Rett变异型患者中，我们没有发现较大的缺失。我们发现，MLPA在寻找危及MECP2整个外显子的大片段缺失时是有用的。作为DNA测序的补充方法，它在经典RS患者中发现了新的MECP2突变。
Mutations in the methyl-CpG-binding protein-2 (MECP2) gene on Xq28 have been found to be a cause of Rett syndrome (RS). In a previous mutation screening, we found MECP2 mutations in 81% of Swedish classical Rett women. In this study, we have analyzed 22 patients for MECP2 deletions using multiplex-ligation-dependent probe amplification (MLPA). Clinically, 11 of the patients who were classical Rett women, 3 were forme fruste, I was congenital RS, and 7 were Rett variants. As inclusion criteria, we used DNA from patients in whom previous sequencing results showed no mutations in coding portions of the MECP2 gene. MLPA is a method based on multiplex PCR. In one PCR, as many as 40 probes are amplified with the same primers. The specificity of the amplification products is determined by the site-specific hybridization of each probe construct, prior to amplification. Each PCR product has a unique length, which makes it possible to identify it by size separation. In 3 of 11 (27%) classical Rett women, we detected large deletions in MECP2 using MLPA. All these patients had deletions covering two exons; in 2 cases the deletion involved exons 3 and 4 and, in one case, exons I and 2 were missing. In the forme fruste, congenital and Rett-variant patients, we found no large deletions. We have found that MLPA is useful when it comes to finding large deletions compromising whole exons in MECP2. Used as a complementary method to DNA sequencing, it revealed new MECP2 mutations in classical RS patients.