Nuclei Isolation From Murine and Human Periosteum For Transcriptomic Analyses
Abstract
Bone repair is a complex regenerative process relying on skeletal stem/progenitor cells (SSPCs) recruited predominantly from the periosteum. Activation and differentiation of periosteal SSPCs occurs in an heterogeneous environment, rising the need of using single cell/nucleus transcriptomics in order to decipher the response of the periosteum to injury. Yet, enzymatic cell dissociation can induce a stress response affecting the transcriptome and lead to overrepresentation of certain cell types (i.e. immune and endothelial cells) and low coverage of other cell types of interest. To counteract these limitations, we optimized a protocol to isolate nuclei directly from the intact periosteum and from the fracture callus to perform single-nucleus RNA sequencing (snRNAseq). This protocol is adapted for fresh murine periosteum, fracture callus and for frozen human periosteum. Nuclei are isolated using mechanical extraction combined with fluorescence-based nuclei sorting to obtain high-quality nucleus suspension. This protocol allows to capture the full diversity of cell types in the periosteum and the fracture environment to better reflect the in vivo tissue composition.
Related
- Fluorescence-activated nuclei sorting (FANS) on human post-mortem cortex tissue enabling the isolation of distinct neural cell populations for multiple omic profiling v2
- Morphological and transcriptomic analyses reveal the embryonic development of Macrobrachium rosenbergii
- Integrated toxicogenomic and single-nucleus transcriptomic analyses prioritize SDHB and NDUFAB1 in mitochondrial respiration-associated remodeling during cisplatin-induced acute kidney injury
- Integrated bulk and single-cell transcriptomic analyses with experimental validation identify PLOD1 as a diagnostic biomarker and TGF-β1-responsive profibrotic mediator in dupuytren’s disease
- Integrated transcriptomic and metabolomic analyses reveal that flavonoid metabolic pathways are associated with petal discoloration in Aquilegia viridiflora s.l.
BibTeX
@article{perrin2026nuclei,
title = {{Nuclei Isolation From Murine and Human Periosteum For Transcriptomic Analyses}},
author = {Simon Perrin and Cassandre Goachet and Maria Ethel and Yasmine Hachemi and Céline Colnot},
journal = {Bio-protocol},
year = {2026},
month = sep,
doi = {10.21769/bioprotoc.5211},
url = {https://u-pec.hal.science/hal-05772616v1},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Nuclei Isolation From Murine and Human Periosteum For Transcriptomic Analyses
AU - Simon Perrin
AU - Cassandre Goachet
AU - Maria Ethel
AU - Yasmine Hachemi
AU - Céline Colnot
PY - 2026
DA - 2026/09/30
JO - Bio-protocol
DO - 10.21769/bioprotoc.5211
UR - https://u-pec.hal.science/hal-05772616v1
AB - Bone repair is a complex regenerative process relying on skeletal stem/progenitor cells (SSPCs) recruited predominantly from the periosteum. Activation and differentiation of periosteal SSPCs occurs in an heterogeneous environment, rising the need of using single cell/nucleus transcriptomics in order to decipher the response of the periosteum to injury. Yet, enzymatic cell dissociation can induce a stress response affecting the transcriptome and lead to overrepresentation of certain cell types (i.e. immune and endothelial cells) and low coverage of other cell types of interest. To counteract these limitations, we optimized a protocol to isolate nuclei directly from the intact periosteum and from the fracture callus to perform single-nucleus RNA sequencing (snRNAseq). This protocol is adapted for fresh murine periosteum, fracture callus and for frozen human periosteum. Nuclei are isolated using mechanical extraction combined with fluorescence-based nuclei sorting to obtain high-quality nucleus suspension. This protocol allows to capture the full diversity of cell types in the periosteum and the fracture environment to better reflect the in vivo tissue composition.
SN - 2331-8325
ER -