Université de Paris, CNRS, Institut Jacques Monod, Paris, France.
Institut Curie, PSL Research University, CNRS UMR 3348, Orsay, France.
Methods Mol Biol. 2022;2477:35-55. doi: 10.1007/978-1-0716-2257-5_3.
Detecting protein-RNA interactions in vivo is essential for deciphering many important cellular pathways. Several methods have been described for this purpose, among which cross-linking analysis of cDNA, CRAC. This method relies on a first step of UV cross-linking of living yeast cells and several subsequent steps of purification of the protein-RNA complexes, some of which under denaturing condition. Without altering the general principle of the method, we have modified and improved the protocol, with the specific aim of sequencing the nascent RNA isolated from transcription complexes and generate high-resolution and directional transcription maps.
在体内检测蛋白质-RNA 相互作用对于破译许多重要的细胞途径至关重要。为此目的已经描述了几种方法，其中包括 cDNA 的交联分析、CRAC。该方法依赖于对活酵母细胞进行紫外线交联的第一步，以及随后的几个纯化蛋白质-RNA 复合物的步骤，其中一些是在变性条件下进行的。在不改变该方法的一般原理的情况下，我们已经对该方案进行了修改和改进，具体目的是对从转录复合物中分离出的新生 RNA 进行测序，并生成高分辨率和定向的转录图谱。
