使用TAT-Cre重组酶在ex vivo精密切割肺切片 (PCLS) 中进行基因操纵的一种新方法
Sek-Shir Cheong1, Tiago C Luis2, Matthew Hind1,3
1National Heart and Lung Institute (NHLI), Imperial College London, London, UK.
Bio-protocol
|April 30, 2024
概括
通过TReATS方法,可以在四天内在精密切割肺切片 (PCLS) 中快速基因修饰. 这种ex vivo方法通过简化遗传研究和减少动物使用来加速肺部研究.
科学领域:
- 肺研究研究 肺研究
- 实体组织模拟的ex vivo组织建模.
- 基因工程是一种基因工程.
背景情况:
- 精密切割肺切片 (PCLS) 是有价值的3D肺模型,保留了原生细胞和细胞外矩阵环境.
- 在PCLS中基因修饰的现有方法可能是复杂和耗时的.
研究的目的:
- 介绍TReATS (组织切片中的TAT-Cre复合酶介导的浮性等位基因修饰) 方法,用于PCLS中快速有效的基因修饰.
- 提供一个简化,可复制的协议,用于肺组织的ex vivo基因操纵.
主要方法:
- 从成年羊动物中产生PCLS.
- 在TAT-Cre重组酶溶液中化PCLS.
- 通过活着染色,成像,qPCR和细胞活力测试进行验证.
主要成果:
- 在为期四天的协议中,PCLS成功进行基因修饰.
- 消除对复杂的养策略的需要.
- 规避与因基因突变导致的过早死亡相关的问题.
结论:
- 在PCLS中,TReATS方法提供了一个简单,快速和可重复的解决方案,用于ex vivo基因修饰.
- 这种技术加速了对基因功能,肺病机制和药物向发现的研究.
- 该方法适用于基因删除或激活,并对其他器官组织切片有潜在的应用.
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