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RNA降解对纳米孔直接RNA测序的普遍影响
Yair D J Prawer1, Josie Gleeson1, Ricardo De Paoli-Iseppi1
1Centre for Stem Cell Systems, Department of Anatomy and Physiology, The University of Melbourne, Parkville, VIC, 3010, Australia.
NAR genomics and bioinformatics
|June 12, 2023
概括
直接RNA测序 (DRS) 可能因RNA降解而产生偏差,影响基因表达测量. 然而,显式校正方法可以恢复生物信号,使得DRS适用于部分降解的样品.
科学领域:
- 分子生物学分子生物学
- 基因组学就是基因组学.
- 生物信息学是一种生物信息学.
背景情况:
- 牛津纳米孔直接RNA测序 (DRS) 提供全长RNA分析和精确的基因表达量化.
- 与其他方法相比,DRS依赖完整的RNA引发了人们对RNA完整性对表达测量的影响的担忧.
- 对于RNA降解对DRS的影响以及纠正的可能性仍然不清楚.
研究的目的:
- 研究RNA降解对直接RNA测序 (DRS) 性能的影响.
- 为了确定DRS中的RNA降解效应是否可以被纠正.
- 制定使用不同RNA完整性的样本DRS的指导方针.
主要方法:
- 使用SH-SY5Y神经母细胞瘤细胞进行了降解时间序列实验.
- 评估了RNA完整性,并在不同降解阶段的样本上进行了DRS.
- 进行了差异表达分析,并没有明确的降解校正.
主要成果:
- RNA降解显著偏差DRS测量,减少库的复杂性和过度代表短基因/异形.
- 降解也会影响差异表达分析,但纠正方法在很大程度上可以恢复生物信号.
- 对于部分降解的样本,DRS显示的偏差比纳米孔PCR-cDNA测序更小.
结论:
- 具有RNA完整数 (RIN) >9.5的样本被认为是DRS.的未降级样本.
- DRS适用于具有RIN> 7的样品,当应用适当的降解校正时.
- DRS适用于各种样品类型,包括临床和死后组织,有效管理降解效应.
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