一个精确而高效的循环RNA合成系统,基于一种来自Tetrahymena thermophila的 ribozyme
Jingyi Cui1,2,3, Lanxin Zhang3, Zaifeng Zhang1,2
1The Key Laboratory of Geriatrics, Beijing Institute of Geriatrics, Institute of Geriatric Medicine, Chinese Academy of Medical Sciences, Beijing Hospital/National Center of Gerontology of National Health Commission, PR China.
Nucleic acids research
|June 28, 2023
概括
这项研究引入了一种基于 ribozyme 的新型系统,用于有效的循环 RNA (circRNA) 制备,避免额外的核酸. 经过工程设计的circRNA保留了生物功能,并且可以被翻译,使未来的功能研究和大规模生产成为可能.
科学领域:
- 分子生物学分子生物学
- 在RNA生物学,RNA生物学.
- 生物技术是生物技术.
背景情况:
- 传统的循环RNA (circRNA) 制备方法通常会导致线性转录或不需要的核酸添加.
- 现有的circRNA合成方法缺乏效率和精度,阻碍了功能研究和应用.
研究的目的:
- 开发一种高效和精确的循环RNA (circRNA) 制备系统,使用自剪接的 ribozyme.
- 为了证明由 ribozyme 生成的 circRNA 的功能相关性及其表达潜力.
主要方法:
- 从一个优化的Tetrahymena thermophila I组内设计了一个自剪接的 ribozyme,用于RNA循环化.
- 插入向RNA序列的目标下游的 ribozyme 和一个补充的反意义区域上游,以促进循环.
- 通过使用特定的基因 (DNMT1,CDR1as,FOXO3,HIPK3) 将 ribozyme 中介循环化的效率与 flanking 内基互补序列 (ICS) 中介方法进行了比较.
- 使用分裂的绿色光蛋白 (GFP) 和Coxsackievirus B3 (CVB3) 内部核糖体进入点 (IRES) 证明了基于核糖酶的循环mRNA表达系统.
主要成果:
- 与侧面的ICS方法相比, ribozyme介导的系统表现出明显更高的循环化效率.
- 通过 ribozyme 介导的循环化产品没有额外的核酸.
- 过度表达的circFOXO3保持了其生物功能,包括调节细胞增殖,迁移和细胞亡.
- 基于 ribozyme 的循环 mRNA 表达系统成功实现了循环 mRNA 的翻译.
结论:
- 开发的基于 ribozyme 的系统提供了一种新,方便和快速的方法来设计RNA循环化.
- 该系统适用于循环RNA的功能研究和大规模制备.
- 产生功能性和可翻译的循环RNA的能力在RNA疗法和研究中开辟了新的途径.
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