PCR amplicons identify widespread copy number variation in human centromeric arrays and instability in cancer
PCR amplicons identify widespread copy number variation in human centromeric arrays and instability in cancer
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PCR 扩增子可识别人类着丝粒阵列中广泛的拷贝数变异和癌症的不稳定性
DOI：
10.1016/j.xgen.2021.100064
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发表时间：
2021-12-08
期刊：
影响因子：
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通讯作者：
Gerton, Jennifer L.
中科院分区：
文献类型：
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作者：
de Lima, Leonardo Gomes;Howe, Edmund;Gerton, Jennifer L.
着丝粒α-卫星重复序列占人类基因组的约6%，但它们的长度和重复性质使这些区域的测序和分析具有挑战性。然而，着丝粒对于染色体的稳定繁殖是必不可少的，因此迫切需要工具来监测着丝粒拷贝数以及它如何影响染色体传递和基因组稳定性。我们开发和基准液滴数字PCR（ddPCR）测定，测量拷贝数的五个人类着丝粒阵列。我们应用它们来表征着丝粒阵列大小的自然变异，分析了来自中国的37个个体和来自美国和英国的39个个体的正常组织。每个染色体特异性阵列的大小在个体之间变化高达10倍，在染色体之间变化高达50倍，这表明每个个体中的阵列具有独特的互补性。我们还使用ddPCR分析了四种癌症类型中76个匹配的肿瘤正常样本中的着丝粒拷贝数，这是迄今为止对癌症中着丝粒阵列稳定性的最全面的定量分析。与培养细胞中的稳定传输相反，着丝粒阵列显示每种癌症类型中的获得和丢失事件，表明着丝粒a-卫星DNA代表癌症中基因组不稳定性的新类别。我们测量人类着丝粒阵列拷贝数的方法将推进正常和疾病状态下着丝粒和基因组完整性的研究。
Centromeric a-satellite repeats represent-6% of the human genome, but their length and repetitive nature make sequencing and analysis of those regions challenging. However, centromeres are essential for the stable propagation of chromosomes, so tools are urgently needed to monitor centromere copy number and how it influences chromosome transmission and genome stability. We developed and benchmarked droplet digital PCR (ddPCR) assays that measure copy number for five human centromeric arrays. We applied them to characterize natural variation in centromeric array size, analyzing normal tissue from 37 individuals from China and 39 individuals from the US and UK. Each chromosome-specific array varies in size up to 10-fold across individuals and up to 50-fold across chromosomes, indicating a unique complement of arrays in each individual. We also used the ddPCR assays to analyze centromere copy number in 76 matched tumor-normal samples across four cancer types, representing the most-comprehensive quantitative analysis of centromeric array stability in cancer to date. In contrast to stable transmission in cultured cells, centromeric arrays show gain and loss events in each of the cancer types, suggesting centromeric a-satellite DNA represents a new category of genome instability in cancer. Our methodology for measuring human centromeric-array copy number will advance research on centromeres and genome integrity in normal and disease states.