Isolating and cryopreserving pig skin cells for single-cell RNA sequencing study

The pig skin architecture and physiology are similar to those of humans. Thus, the pig model is very valuable for studying skin biology and testing therapeutics. The single-cell RNA sequencing (scRNA-seq) technology allows quantitatively analyzing cell types, compositions, states, signaling, and rec...

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Bibliographic Details
Published in:PloS one 2022-02, Vol.17 (2), p.e0263869-e0263869
Main Authors: Han, Li, Jara, Carlos P, Wang, Ou, Shi, Yu, Wu, Xinran, Thibivilliers, Sandra, Wóycicki, Rafał K, Carlson, Mark A, Velander, William H, Araújo, Eliana P, Libault, Marc, Zhang, Chi, Lei, Yuguo
Format: Article
Language:English
Subjects:
Analysis
Animals
Biology and Life Sciences
Biomedical engineering
Cell aggregation
Cell culture
Cell death
Cell viability
Cryopreservation
Cryopreservation - methods
Cytotoxicity
Dissociation
Engineering
Enzymes
Exome Sequencing
Gene expression
Gene Expression Profiling
Gene sequencing
Genetic aspects
Hogs
Horticulture
Laboratories
Life sciences
Medicine and Health Sciences
Nursing schools
Pigskins
Plant sciences
Research and Analysis Methods
Ribonucleic acid
RNA
RNA sequencing
Single-Cell Analysis - methods
Skin
Skin - cytology
Skin - metabolism
Skin tests
Specimen Handling - methods
Swine
Transcriptome
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Summary:The pig skin architecture and physiology are similar to those of humans. Thus, the pig model is very valuable for studying skin biology and testing therapeutics. The single-cell RNA sequencing (scRNA-seq) technology allows quantitatively analyzing cell types, compositions, states, signaling, and receptor-ligand interactome at single-cell resolution and at high throughput. scRNA-seq has been used to study mouse and human skins. However, studying pig skin with scRNA-seq is still rare. A critical step for successful scRNA-seq is to obtain high-quality single cells from the pig skin tissue. Here we report a robust method for isolating and cryopreserving pig skin single cells for scRNA-seq. We showed that pig skin could be efficiently dissociated into single cells with high cell viability using the Miltenyi Human Whole Skin Dissociation kit and the Miltenyi gentleMACS Dissociator. Furthermore, the obtained single cells could be cryopreserved using 90% FBS + 10% DMSO without causing additional cell death, cell aggregation, or changes in gene expression profiles. Using the developed protocol, we were able to identify all the major skin cell types. The protocol and results from this study are valuable for the skin research scientific community.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0263869