Protocol for CRISPR-Cas9 modification of glycosylation in 3D organotypic skin models

Irina N Marinova; Hans H Wandall; Sally Dabelsteen

doi:10.1016/j.xpro.2021.100668

Protocol for CRISPR-Cas9 modification of glycosylation in 3D organotypic skin models

STAR Protoc. 2021 Jul 10;2(3):100668. doi: 10.1016/j.xpro.2021.100668. eCollection 2021 Sep 17.

Authors

Irina N Marinova¹, Hans H Wandall¹, Sally Dabelsteen²

Affiliations

¹ Copenhagen Center for Glycomics, Department of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.
² Department of Oral Pathology, School of Dentistry, University of Copenhagen, Copenhagen, Denmark.

Abstract

Glycosylation is one of the most common protein modifications in living organisms and has important regulatory roles in animal tissue development and homeostasis. Here, we present a protocol for generation of 3D organotypic skin models using CRISPR-Cas9 genetically engineered human keratinocytes (N/TERT-1) to study the role of glycans in epithelial tissue formation. This strategy is also applicable to other gene targets and organotypic tissue models. Careful handling of the cell cultures is critical for the successful formation of the organoids. For complete details on the use and execution of this protocol, please refer to Dabelsteen et al. (2020).

Keywords: CRISPR; Cell Biology; Organoids.

Publication types

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

MeSH terms

CRISPR-Cas Systems*
Fibroblasts
Gene Knockout Techniques / methods*
Glycosylation
HEK293 Cells
Humans
Keratinocytes / cytology
Lentivirus / genetics
Organoids / cytology*
Organoids / physiology
Skin / cytology*