Large genomic mutations within the ATM gene detected by MLPA, including a duplication of 41 kb from exon 4 to 20
Large genomic mutations within the ATM gene detected by MLPA, including a duplication of 41 kb from exon 4 to 20
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DOI：
10.1111/j.1469-1809.2007.00399.x
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发表时间：
2008-01-01
影响因子：
1.9
通讯作者：
Brusco, Alfredo
中科院分区：
文献类型：
--
作者：
Cavalieri, Simona;Funaro, Ada;Brusco, Alfredo
突变检测仍然是大基因的问题，主要是因为基于PCR的方法无法检测杂合缺失和任何重复。迄今为止，在ATM基因中，只描述了少数多外显子缺失，并且这种类型的突变被认为是罕见的。为了解决这个问题，我们测试了一种新的MLPA（多重连接探针扩增）试剂盒，该试剂盒覆盖了66个ATM外显子中的33个，除了3名A-T患者外，还使用了两个先前表征的基因组缺失作为对照，这些患者来自对9名患者的调查，这些患者在常规突变筛查后缺失了4个未识别的突变。我们首次发现：1）一个类似于41 kb的基因组重复跨越外显子4-20（c. 30_2816dup41kb）（又称，ATM dup 41 kb）; 2）包括外显子31的新的基因组缺失，和3）在半合子病中，非缺失外显子31中的点突变。在这项研究中，我们扩展了突变检测9个新的意大利A-T患者，使用单倍型分析，DHPLC和MLPA的组合方法。总体而言，我们实现了> 97%的突变检测率，并且现在可以基于21个连续的意大利A-T家族来定义ATM突变谱。
Mutation detection remains problematic for large genes, primarily because PCR-based methodology fails to detect heterozygous deletions and any duplication. In the ATM gene only a handful of multi-exon deletions have been described to date, and this type of mutation has been considered rare. To address this issue we tested a new MLPA (Multiplex Ligation Probe Amplification) kit that covers 33 of the 66 ATM exons, using for controls two previously characterized genomic deletions in addition to three A-T patients, taken from a survey of nine, who had missing four mutations unidentified after conventional mutation screening. We identified for the first time: 1) a similar to 41 kb genomic duplication spanning exons 4-20 (c.-30_2816dup41kb)(a.k.a., ATM dup 41 kb); 2) a novel genomic deletion including exon 31, and 3) in hemizygosis a point mutation in the non-deleted exon 31. In this study we extended mutation detection to nine new Italian A-T patients, using a combined approach of haplotype analysis, DHPLC and MLPA. Overall we achieved a mutation detection rate of > 97%, and can now define a spectrum of ATM mutations based on twenty-one consecutive Italian families with A-T.