Single-Cell Isolation from Regenerating Murine Muscles for RNA-Sequencing Analysis

Area of Science:

• Molecular Biology
• Genomics
• Regenerative Medicine

Background:

• Single-cell RNA sequencing (scRNA-seq) enables high-resolution analysis of cellular heterogeneity.
• Skeletal muscle regeneration involves complex cellular dynamics.
• Existing protocols may require adaptation for specific tissue types like skeletal muscle.

Purpose of the Study:

• To present a comprehensive protocol for unbiased evaluation of regenerating murine skeletal muscle using scRNA-seq.
• To focus on the isolation of mononuclear cells from skeletal muscle tissue.
• To provide a adaptable method for assessing cell populations in various tissues.

Main Methods:

• Isolation of mononuclear cells from murine skeletal muscle.
• Application of single-cell RNA sequencing (scRNA-seq) for gene expression analysis.
• Unbiased evaluation of cellular composition in regenerating muscle.

Main Results:

• The protocol allows for the deconvolution and clustering of thousands of cells based on gene expression.
• Successful isolation of mononuclear cells from skeletal muscle tissue was achieved.
• The method provides a foundation for detailed analysis of muscle regeneration.

Conclusions:

• The presented scRNA-seq protocol offers a robust method for studying skeletal muscle regeneration.
• The protocol's focus on mononuclear cells is crucial for understanding muscle-specific cellular responses.
• This method can be adapted for scRNA-seq analysis in other complex tissues.