一个复活的Cas12a的祖先扩展了目标访问和基质识别,用于核酸编辑和检测
Ylenia Jabalera1, Igor Tascón2,3, Sara Samperio1
1Center for Cooperative Research in Biosciences (CIC bioGUNE), Basque Research and Technology Alliance (BRTA), Derio, Spain.
Nature biotechnology
|November 1, 2024
概括
祖先序列重建 (ASR) 创造了ReChb,一个Cas12a核酶变体. 在人类细胞中,ReChb提供了几乎没有PAM的基因组编辑和广泛的核酸激活,扩大了CRISPR技术的应用.
科学领域:
- 基因编辑技术CRISPR是基因编辑技术.
- 分子生物学分子生物学
- 结构生物学是结构生物学.
背景情况:
- Cas12a核酶在目标可访问性和应用方面存在局限性.
- 天然Cas12a的 ортолог具有特定的PAM要求和目标识别配置文件.
研究的目的:
- 通过使用祖先序列重建 (ASR) 来设计一种具有扩展准能力的新型Cas12a变体.
- 描述重建的Cas12a祖先的生物化学和结构性质,命名为ReChb.
主要方法:
- 从水生细菌获得的Cas12a正方体的祖先序列重建 (ASR).
- 在人类细胞中进行基因组编辑实验.
- 低温电子显微镜 (cryo-EM) 用于结构分析.
主要成果:
- ReChb表现出接近PAM的无向定位,使得以前无法访问的位置能够进行基因组编辑.
- ReChb可以被各种核酸点激活,包括单链DNA和RNA.
- ReChb证明了DNA和RNA基质的高效附带裂解.
- 化EM揭示了ReChb增强生物物理活动的结构基础.
结论:
- ReChb显著扩大了Cas12a核酶的准范围和多功能性.
- ASR是一种强大的方法来增强CRISPR技术.
- 该ReChb变异对各种基因组工程应用具有前景.
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