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用化α-螺旋聚在体外和体内进行siRNA介导的U1 snRNA淘汰的协议
Fan Zhang1, Chenglong Ge2, Zigang Qiao1
1National Key Laboratory of Immunity and Inflammation, and CAMS Key Laboratory of Synthetic Biology Regulatory Elements, Suzhou Institute of Systems Medicine, Chinese Academy of Medical Sciences & Peking Union Medical College, Suzhou 215123, China.
STAR protocols
|August 3, 2024
概括
这项研究引入了一种可靠的方法,用于降低U1小核RNA (snRNA) 水平,使用由化聚传递的小干扰RNA (siRNA). 该协议有效地使细胞和小鼠肺中的U1 snRNA沉默,有助于研究其功能.
科学领域:
- 分子生物学分子生物学
- 在RNA干扰过程中,RNA干扰
- 生物化学 生物化学
背景情况:
- U1小核RNA (snRNA) 在mRNA前拼接中起着关键作用.
- U1 snRNA的失调与各种病理生理条件有关.
- 对于功能性研究,需要有效的向U1 snRNA淘汰方法.
研究的目的:
- 为了在体外和体内呈现一个强大的U1 snRNA淘汰协议.
- 为了证明化α螺旋多 (P7F7) 对siRNA传递的有效性.
- 为了使U1 snRNA的生理和病理生理作用的调查.
主要方法:
- 准备P7F7/siRNA多复合体进行向传递.
- 在巨细胞培养物中应用多复合体对U1 snRNA沉默.
- 在体内向小鼠肺部注射多重复合物以使基因沉默.
- 使用逆转录定量实时PCR (RT-qPCR) 验证淘汰效率.
主要成果:
- 成功制备了P7F7/siRNA多复合体.
- 使用开发的协议,有效地静止巨细胞中的U1 snRNA.
- 在小鼠肺部体内证明了U1 snRNA的敲除.
- RT-qPCR证实了U1 snRNA水平的显著降低.
结论:
- 本方案提供了一种可靠的方法,用于U1 snRNA的淘汰.
- 化聚胺介导的siRNA传递在细胞和动物模型中对U1 snRNA沉默有效.
- 这种方法有助于研究U1 snRNA在健康和疾病中的功能.
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