Extraction and sequencing of single nuclei from murine skeletal muscles

Matthieu Dos Santos; Stamatia Gioftsidi; Stéphanie Backer; Léo Machado; Frederic Relaix; Pascal Maire; Philippos Mourikis

doi:10.1016/j.xpro.2021.100694

Extraction and sequencing of single nuclei from murine skeletal muscles

STAR Protoc. 2021 Jul 28;2(3):100694. doi: 10.1016/j.xpro.2021.100694. eCollection 2021 Sep 17.

Authors

Matthieu Dos Santos¹, Stamatia Gioftsidi², Stéphanie Backer¹, Léo Machado², Frederic Relaix^{2

3

4

5}, Pascal Maire¹, Philippos Mourikis²

Affiliations

¹ Université de Paris, Institut Cochin, INSERM, CNRS, 75014 Paris, France.
² Univ Paris Est Créteil, INSERM, IMRB, 94010 Créteil, France.
³ EnvA, IMRB, 94700 Maisons-Alfort, France.
⁴ EFS, IMRB, 94010 Creteil, France.
⁵ AP-HP, Hopital Mondor, Service d'histologie, 94010 Creteil, France.

Abstract

Single-nucleus RNA sequencing allows the profiling of gene expression in isolated nuclei. Here, we describe a step-by-step protocol optimized for adult mouse skeletal muscles. This protocol provides two main advantages compared to the widely used single-cell protocol. First, it allows us to sequence the myonuclei of the multinucleated myofibers. Second, it circumvents the cell-dissociation-induced transcriptional modifications. For complete details on the use and execution of this protocol, please refer to Dos Santos et al. (2020) and Machado, Geara et al. (2021).

Keywords: Cell Differentiation; Cell isolation; Flow Cytometry/Mass Cytometry; RNAseq; Single Cell; Stem Cells.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Cell Culture Techniques / methods
Cell Nucleus / genetics*
Cell Separation / methods
Mice
Muscle Fibers, Skeletal / metabolism
Muscle, Skeletal / cytology
RNA / chemistry
RNA / isolation & purification
Satellite Cells, Skeletal Muscle / metabolism
Sequence Analysis, RNA / methods*
Single-Cell Analysis / methods*

Substances

RNA