利用高效自催化转录本连接 RNA 以生成杂交环状 RNA。
Trans ligation of RNAs to generate hybrid circular RNAs using highly efficient autocatalytic transcripts.
机构信息
Department of Pharmacology, Weill-Cornell Medical College, Cornell University, New York, NY 10065, USA.
Department of Pharmacology, Weill-Cornell Medical College, Cornell University, New York, NY 10065, USA.
出版信息
Methods. 2021 Dec;196:104-112. doi: 10.1016/j.ymeth.2021.05.009. Epub 2021 May 13.
Circular RNAs are useful entities for various biotechnology applications, such as templating translation and binding or sequestering miRNA and RNA binding proteins. Circular RNA as highly resistant to degradation in cells and are more long-lived than linear RNAs. Here, we describe a method for intracellular trans ligation of RNA transcripts that can generate hybrid circular RNAs. These hybrid circular RNAs comprise two separate RNA that are covalently linked by ligation to form a circular RNA. By incorporating self-cleaving ribozymes at each site of ligation, trans ligation of the transcripts occurs in mammalian cells with no additional material. We provide a protocol for designing and testing trans ligation of transcripts and demonstrate detection of hybrid circular RNAs using fluorescence microscopy.
环状 RNA 是各种生物技术应用的有用实体，例如模板翻译和结合或隔离 miRNA 和 RNA 结合蛋白。环状 RNA 在细胞中具有很强的抗降解能力，比线性 RNA 寿命更长。在这里，我们描述了一种用于细胞内 RNA 转录物连接的方法，该方法可以产生杂交环状 RNA。这些杂交环状 RNA 由两个单独的 RNA 通过连接共价连接而成，形成环状 RNA。通过在每个连接位点掺入自我切割核酶，在没有额外物质的情况下，在哺乳动物细胞中发生转录物的连接。我们提供了一种设计和测试转录物连接的方案，并通过荧光显微镜显示了杂交环状 RNA 的检测。
Zotero 插件