Making an unbiased library Sequencing the genome of single cells gives insight into issues such as cell-to-cell heterogeneity and genome instability. Key to single-cell sequencing techniques are whole-genome amplification (WGA) methods that provide sufficient DNA for next-generation sequencing. Current WGA methods have been hampered by low accuracy and spatial resolution of gene copy numbers and by low amplification fidelity. Chen et al. report an improved single-cell WGA method, Linear Amplification via Transposon Insertion (LIANTI). The DNA is randomly fragmented by Tn5 transposition of a transposon that includes a T7 promoter, which allows linear amplification. The authors used the method to determine the spectrum of single-nucleotide variations in a single human cell after ultraviolet radiation. Science, this issue p. 189 An improved whole-genome amplification method reduces amplification bias and errors. Single-cell genomics is important for biology and medicine. However, current whole-genome amplification (WGA) methods are limited by low accuracy of copy-number variation (CNV) detection and low amplification fidelity. Here we report an improved single-cell WGA method, Linear Amplification via Transposon Insertion (LIANTI), which outperforms existing methods, enabling micro-CNV detection with kilobase resolution. This allowed direct observation of stochastic firing of DNA replication origins, which differs from cell to cell. We also show that the predominant cytosine-to-thymine mutations observed in single-cell genomics often arise from the artifact of cytosine deamination upon cell lysis. However, identifying single-nucleotide variations (SNVs) can be accomplished by sequencing kindred cells. We determined the spectrum of SNVs in a single human cell after ultraviolet radiation, revealing their nonrandom genome-wide distribution.

中文翻译：

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通过转座子插入线性扩增 (LIANTI) 进行单细胞全基因组分析

制作无偏文库 对单细胞基因组进行测序可以深入了解细胞间异质性和基因组不稳定性等问题。单细胞测序技术的关键是全基因组扩增 (WGA) 方法，可为下一代测序提供足够的 DNA。当前的 WGA 方法受到基因拷贝数的低准确性和空间分辨率以及低扩增保真度的阻碍。陈等人。报告了一种改进的单细胞 WGA 方法，通过转座子插入的线性放大 (LIANTI)。DNA 被包含 T7 启动子的转座子的 Tn5 转座随机片段化，从而允许线性扩增。作者使用该方法确定了紫外线辐射后单个人类细胞中单核苷酸变异的光谱。科学，这个问题 p。189 改进的全基因组扩增方法可减少扩增偏差和错误。单细胞基因组学对生物学和医学很重要。然而，当前的全基因组扩增 (WGA) 方法受到拷贝数变异 (CNV) 检测准确性低和扩增保真度低的限制。在这里，我们报告了一种改进的单细胞 WGA 方法，通过转座子插入的线性放大 (LIANTI)，它优于现有方法，能够以千碱基分辨率进行微 CNV 检测。这允许直接观察 DNA 复制起点的随机激发，这在细胞与细胞之间是不同的。我们还表明，在单细胞基因组学中观察到的主要胞嘧啶到胸腺嘧啶突变通常是由细胞裂解时胞嘧啶脱氨的假象引起的。然而，鉴定单核苷酸变异 (SNV) 可以通过对同类细胞进行测序来完成。我们确定了紫外线辐射后单个人类细胞中 SNV 的光谱，揭示了它们的非随机全基因组分布。