Approaches for single-cell RNA sequencing across tissues and cell types

Area of Science:

• Molecular Biology
• Genomics
• Cell Biology

Background:

• Single-cell RNA sequencing (scRNA-seq) has revolutionized the study of cellular heterogeneity and signaling in development and disease.
• Numerous complementary assays exist for profiling individual cell transcriptomes, often combined with other data types like chromatin accessibility or surface protein markers.
• The rapid advancement of scRNA-seq technologies presents challenges in establishing robust, up-to-date protocols for diverse research questions.

Approach:

• This review examines various scRNA-seq techniques, from mRNA end-counting to total RNA profiling, detailing their specific features.
• It outlines essential sample preparation steps and quality control measures crucial for successful single-cell experiments.
• Guidance is provided on automating workflows and managing the expanding landscape of scRNA-seq techniques, drawing from experience in a central single-cell facility.

Key Points:

• Understanding the spectrum of scRNA-seq methods (e.g., 3' end, full-length) and their applications is critical.
• Standardized sample preparation and rigorous quality control are paramount for reliable scRNA-seq data.
• Workflow automation and strategic protocol selection are key to efficiently navigating the expanding scRNA-seq technology portfolio.

Conclusions:

• This review offers practical insights for researchers and core facilities to optimize scRNA-seq workflows.
• It aims to facilitate the effective application of diverse scRNA-seq techniques to address complex biological questions.
• By providing guidance on protocol selection and automation, this work supports robust cellular and molecular analyses.