在使用Hawk-SeqTM的TK6细胞中由变异原体诱导的全基因组突变分析
Yuki Otsubo1, Takako Hirose1, Shoji Matsumura1
1R&D -Safety Science Research, Kao Corporation, 3-25-14 Tono-machi, Kawasaki-ku, Kawasaki-shi, Kanagawa 210-0821, Japan.
概括
使用Hawk-SeqTM的错误纠正下一代测序 (ecNGS) 有效地识别了人类细胞中化学诱导的突变. 这种方法显著降低了背景噪声,改善了罕见遗传变化的检测.
科学领域:
- 基因组学就是基因组学.
- 毒理学 毒理学 毒理学
- 分子生物学分子生物学
背景情况:
- 错误纠正下一代测序 (ecNGS) 对于检测罕见突变至关重要.
- 克隆和亚克隆变异 (CVs和SCVs) 可以降低测序研究中的突变检测灵敏度.
- 人类TK6淋巴细胞细胞系是基因毒性测试的标准模型.
研究的目的:
- 应用Hawk-SeqTM来评估TK6细胞中化学诱导的突变.
- 识别和过出克隆和亚克隆变异,以提高突变检测灵敏度.
- 建立一种可靠的方法来评估MNU和ENU等化学品的变异性.
主要方法:
- 对TK6基因组进行重新测序,以确定4,501,430个克隆变异 (CV).
- 在车辆控制器中过常见基替代 (BS) 以识别和删除亚克隆变异 (SCV).
- 在严格的背景过后,Hawk-SeqTM应用于用N-甲基-N-氨酸urea (MNU) 和N-乙基-N-氨酸urea (ENU) 处理的TK6细胞.
主要成果:
- 车辆控制中的初始BS频率为2.0 × 10-6 bp,在过CV和SCV后降至0.65 × 10-6 bp.
- MNU治疗导致BS频率为9.0 × 10-6 bp,主要是G:C > A:T突变.
- ENU治疗显示BS频率为2.0 × 10-6 bp,G:C > A:T,A:T > C:G和A:T > G:C突变增加.
结论:
- 使用Hawk-SeqTM与CV/SCV过开发的方法显著提高了用于突变检测的信号噪声比.
- 该方法准确地确定了MNU和ENU的独特突变性特征,反映了它们已知的机制.
- 这种方法有可能在毒理学研究中对突变进行敏感检测,并阐明各种突变性机制.
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