MERR APEX-seq protocol for profiling the subcellular nascent transcriptome in mammalian cells.

Area of Science:

• Molecular Biology
• Cell Biology
• Genomics

Background:

• Understanding RNA spatial organization is key to deciphering cellular functions.
• Existing methods may lack the spatiotemporal resolution needed for detailed transcriptome mapping.

Purpose of the Study:

• To present a detailed MERR APEX-seq protocol for spatiotemporally resolved mapping of the subcellular transcriptome.
• To enable precise analysis of RNA localization within cultured mammalian cells.

Main Methods:

• Development and description of the MERR APEX-seq protocol.
• Stable cell line construction expressing APEX2.
• MERR APEX labeling, biotinylated RNA enrichment, library preparation, and high-throughput sequencing.
• Detailed data analysis pipeline for MERR APEX-seq.

Main Results:

• The protocol allows for detailed mapping of the subcellular transcriptome with high spatiotemporal resolution.
• Successful application in cultured mammalian cells.

Conclusions:

• The MERR APEX-seq protocol is a valuable tool for investigating RNA spatial organization.
• This technique advances the understanding of RNA functions within the cellular context.