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针对STR类型的CRISPR-Cas9向纳米孔测序

Ting-Ting Yang^1,2,3, Jia-Rong Zhang^1,2,3, Zi-Han Xie^2,4

¹School of Forensic Medicine, Shanxi Medical University, Taiyuan, People's Republic of China.

Electrophoresis

|November 6, 2025

概括

针对CRISPR-Cas9的测序 (Cas9-seq) 为法医分析提供了无PCR的DNA丰富. 然而,这项试点研究发现Cas9-seq并没有改善等位基平衡或减少误差,而不是基于放大方法的短期并联重复造型.

科学领域:

法医遗传学法医遗传学
分子生物学分子生物学
基因组学就是基因组学.

背景情况:

通过CRISPR-Cas9技术,通过单导向RNA (sgRNA) 实现了针对性的DNA丰富.
无聚合酶连锁反应 (PCR) 的工作流程对于法医DNA分析是可取的.
短串重复 (STR) 分析是法医遗传学的基石.

研究的目的:

为了评估CRISPR-Cas9向纳米孔序列 (Cas9-seq) 的有效性,用于法医STR分析.
为了比较Cas9-seq的性能与传统的基于放大功率的纳米孔测序 (amplicon-seq).

主要方法:

开发了一种Cas9-seq方法,针对七个法医STR位置.
使用了人类细胞系DNA (NA12878和293T),输入量为3μg.
将Cas9-seq结果与来自ForenSeq DNA签名准备套件的amplicon-seq进行比较.

主要成果:

在目标地区实现了显著的丰富比率 (643.45x和468.34x).
Cas9-seq表现出超低链偏差,但显示出更高的噪声,并且在等位基因平衡中没有优势.
两种方法都产生了每个样本的3个基因型错误;Cas9-seq没有引入假阳性SNP,与amplicon-seq.不同.

结论:

没有PCR的Cas9-seq显示了高缩和低链偏差.
Cas9-seq在法医STR基因型准确性方面没有超过基于放大功能的方法.
需要进一步优化,以便Cas9-seq被认为有利于法医应用.

关键词:

克里斯普尔-卡斯9是什么意思在前面的Seqq.法医短串重复 (STR) 分析纳米孔测序的测序没有聚合酶连锁反应 (PCR) 的.

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