A protocol for time-resolved transcriptomics through metabolic labeling and combinatorial indexing

Abstract

Summary: The snapshot nature of single-cell transcriptomics presents a challenge for studying the dynamics of gene expression. Metabolic labeling, where nascent RNA is labeled with 4-thiouridine (4sU), captures temporal information at the single-cell level, providing greater insight into expression dynamics. Here, we present an optimized, automation-friendly protocol for the metabolic labeling of RNA alongside single-cell RNA sequencing through combinatorial indexing. We describe steps for 4sU labeling, cell fixation and chemical treatment, and automated two-level combinatorial indexing.For complete details on the use and execution of this protocol, please refer to Maizels et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Keywords

• Cell culture
• Single Cell
• Developmental biology
• RNAseq
• Molecular Biology
• Gene Expression