原生DNA的直接转换用于敏感的多式单分子测序
Arjun S Nanda1,2, Ke Wu1, Iryna Irkliyenko1
1Gladstone Institute for Data Science and Biotechnology, Gladstone Institutes, San Francisco, CA, USA.
Nature genetics
|May 9, 2024
概括
我们开发了标记方法,SMRT-Tag和SAMOSA-Tag,以显著减少PacBio单分子测序的DNA输入. 这些技术可以对遗传变异,DNA甲基化和染色质可访问性进行敏感检测.
科学领域:
- 基因组学就是基因组学.
- 分子生物学分子生物学
- 表观遗传学 在表观遗传学中,表观遗传学是指表观遗传学.
背景情况:
- 太平洋生物科学 (PacBio) 长DNA模板的单分子测序需要大量的输入材料.
- 目前的方法限制了PacBio测序的应用,用于DNA数量有限的研究.
研究的目的:
- 为PacBio测序开发基于标记的新方法,大大减少DNA输入要求.
- 为了实现多式单分子基因组学,包括遗传变异,DNA甲基化和染色质可访问性分析.
主要方法:
- 适应Tn5转移到DNA片段,并引入毛寡核酸来产生PacBio兼容的圆形分子.
- 通过标记 (SMRT-Tag) 开发单分子实时测序,用于基因变异和CpG甲基化检测.
- 开发单分子氨酸甲基化寡核细胞体序列测定试验通过标记 (SAMOSA-Tag) 用于使用外源性氨酸甲基化进行染色体可访问性分析.
主要成果:
- 与现有协议相比,SMRT-Tag和SAMOSA-Tag使用的输入DNA减少了90-99%.
- 使用40ng人类DNA的SMRT-Tag产生了与黄金标准全基因组和双硫酸盐测序相提并论的结果.
- SAMOSA-Tag解决了前列腺癌异种移植中的单纤维色素结构,CTCF结合和DNA甲基化,揭示了与转移相关的表观基因组失调.
结论:
- 基于标记的方法显著降低了PacBio单分子测序的DNA输入要求.
- 这些新型测试有助于敏感,可扩展和多式单分子基因组学.
- 开发的技术有望用于各种基础研究和临床应用.
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