Peking-Tsinghua Center for Life Sciences, Academy for Advanced Interdisciplinary Studies, College of Chemistry and Molecular Engineering, Synthetic and Functional Biomolecules Center Beijing National Laboratory for Molecular Sciences, Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, PKU-IDG/McGovern Institute for Brain Research, Peking University, Beijing 100871, China; Chinese Institute for Brain Research (CIBR), Beijing 102206, China.
STAR Protoc. 2023 Mar 17;4(1):102057. doi: 10.1016/j.xpro.2023.102057. Epub 2023 Jan 24.
Knowledge about the spatial organization of RNAs in eukaryotic cells is crucial for understanding their functions. Here, we present a detailed MERR APEX-seq protocol to achieve spatiotemporally resolved mapping of the subcellular transcriptome in cultured mammalian cells. This protocol provides detailed description of constructing cell lines stably expressing APEX2, immunofluorescence characterization, MERR APEX labeling, enrichment of biotinylated RNA, library construction and high-throughput sequencing, and MERR APEX-seq data analysis. For complete details on the use and execution of this protocol, please refer to Li et al. (2022)..
关于真核细胞中 RNA 空间组织的知识对于理解它们的功能至关重要。在这里，我们提出了一个详细的 MERR APEX-seq 方案，以实现培养的哺乳动物细胞中细胞内转录组的时空分辨作图。该方案提供了构建稳定表达 APEX2 的细胞系、免疫荧光特征、MERR APEX 标记、生物素化 RNA 富集、文库构建和高通量测序以及 MERR APEX-seq 数据分析的详细描述。有关此方案使用和执行的完整详细信息，请参阅 Li 等人（2022）。
