Little is known about the mutational impact of ionizing radiation (IR) exposure on a genome-wide level in mammalian tissues. Recent advancements in sequencing technology have provided powerful tools to perform exome-wide analyses of genetic variation. This also opened up new avenues for studying and characterizing global genomic IR-induced effects. However, genotypes generated by next generation sequencing (NGS) studies can contain errors, which may significantly impact the power to detect signals in common and rare variant analyses. These genotyping errors are not explicitly detected by the standard Genotype Analysis ToolKit (GATK) and Variant Quality Score Recalibration (VQSR) tool and thus remain a potential source of false-positive variants in whole exome sequencing (WES) datasets. In this context, the transition-transversion ratio (Ti/Tv) is commonly used as an additional quality check. In case of IR experiments, this is problematic when Ti/Tv itself might be influenced by IR treatment. It was the aim of this study to determine a suitable threshold for variant filters for NGS datasets from irradiated cells in order to achieve high data quality using Ti/Tv, while at the same time being able to investigate radiation-specific effects on the Ti/Tv ratio for different radiation doses. By testing a variety of filter settings and comparing the obtained results with publicly available datasets, we observe that a coverage filter setting of depth (DP) 3 and genotype quality (GQ) 20 is sufficient for high quality single nucleotide variants (SNVs) calling in an analysis combining GATK and VSQR and that Ti/Tv values are a consistent and useful indicator for data quality assessment for all tested NGS platforms. Furthermore, we report a reduction in Ti/Tv in IR-induced mutations in primary human gingiva fibroblasts (HGFs), which points to an elevated proportion of transversions among IR-induced SNVs and thus might imply that mismatch repair (MMR) plays a role in the cellular damage response to IR-induced DNA lesions.

中文翻译：

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电离辐射改变了人类牙龈成纤维细胞外显子的转移/转化率。

关于电离辐射（IR）暴露对哺乳动物组织中全基因组水平的突变影响知之甚少。测序技术的最新进展提供了强大的工具，可以对基因变异进行全基因组分析。这也为研究和表征全球基因组红外诱导效应开辟了新途径。但是，下一代测序（NGS）研究产生的基因型可能包含错误，这可能会显着影响常见和罕见变体分析中检测信号的能力。标准基因型分析工具包（GATK）和变异质量得分重新校准（VQSR）工具未明确检测到这些基因分型错误，因此在整个外显子组测序（WES）数据集中仍然是假阳性变异的潜在来源。在这种情况下，转换比（Ti / Tv）通常用作附加质量检查。在进行IR实验的情况下，当Ti / Tv本身可能受到IR处理的影响时，这是有问题的。这项研究的目的是确定来自辐照细胞的NGS数据集的变体过滤器的合适阈值，以便使用Ti / Tv获得高数据质量，同时能够研究特定辐射对Ti / Tv的影响。不同辐射剂量的电视比率。通过测试各种过滤器设置并将获得的结果与可公开获得的数据集进行比较，我们观察到深度（DP）3和基因型质量（GQ）20的覆盖率过滤器设置足以满足高质量单核苷酸变体（SNV）的要求，这需要结合GATK和VSQR进行分析，并且Ti / Tv值是一致且有用的所有经过测试的NGS平台的数据质量评估指标。此外，我们报道了原代人牙龈成纤维细胞（HGF）的IR诱导突变中的Ti / Tv降低，这表明IR诱导的SNV中转化的比例增加，因此可能暗示错配修复（MMR）起作用对IR诱导的DNA损伤的细胞损伤反应。