Miniaturization of Smart-seq2 for Single-Cell and Single-Nucleus RNA Sequencing

Area of Science:

• Molecular Biology
• Genomics
• Neuroscience

Background:

• Single-cell RNA sequencing (scRNA-seq) is crucial for understanding cellular heterogeneity.
• Existing protocols can be costly and labor-intensive for large-scale studies.
• Smart-seq2 offers a robust method for full-length transcript analysis.

Purpose of the Study:

• To present a miniaturized, plate-based workflow for Smart-seq2.
• To enable high-throughput, cost-effective single-cell/nucleus RNA sequencing.
• To provide a reliable protocol for mouse and human sample preparation.

Main Methods:

• Cell/nucleus isolation from mouse brain and human organoids.
• Flow sorting of single cells/nuclei into 384-well plates.
• Miniaturized Smart-seq2 library preparation using a liquid-handling robot.

Main Results:

• A reliable and high-throughput protocol for scRNA-seq.
• Cost-effective preparation of samples for deep sequencing.
• Applicability to both mouse and human samples.

Conclusions:

• The described workflow facilitates efficient and scalable full-length single-cell/nucleus RNA sequencing.
• This protocol supports comprehensive transcriptomic analysis in diverse biological contexts.
• The method is optimized for reliability and cost-effectiveness in research settings.