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Tagging and Fusion Proteins

Tagging and Fusion Proteins

Proteins are involved in several cellular processes and biochemical reactions. Analyzing a specific protein of interest requires it to be isolated from the other proteins in the cell. This is achieved by overexpressing the specific gene in a suitable host to produce large quantities of the target protein. A tag or label is recombined with the gene to produce a fusion protein containing the target protein and the tag. The tags on these fusion proteins can then be used for easy detection and...

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Genetic Barcoding with Fluorescent Proteins for Multiplexed Applications

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Published on: April 14, 2015

Multiplexed genetic tags for electron and fluorescence microscopy.

Oleksandr Berezin^1,2, Alberto Piovesan^1,2, Roman Graf^1,2

¹Munich Institute of Biomedical Engineering, Department of Bioscience, TUM School of Natural Sciences and TUM School of Medicine, Technical University of Munich, Munich, Germany.

Nature Protocols

|November 19, 2025

Summary

This protocol details using genetically encoded EMcapsulin reporters for fluorescence and electron microscopy (EM). These modular gene reporters offer distinct shapes for cellular and protein labeling in various research applications.

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Area of Science:

Cell biology
Microscopy techniques
Molecular biology

Background:

Genetically encoded reporters are crucial for visualizing cellular processes.
Electron microscopy (EM) requires specific labeling for high-resolution imaging.
Existing EM labeling methods can be complex and time-consuming.

Purpose of the Study:

To provide a comprehensive protocol for using genetically encoded, multichannel EMcapsulin reporters.
To enable researchers, including those with limited EM experience, to utilize these reporters effectively.
To facilitate the labeling of cells and proteins for both fluorescence microscopy and EM workflows.

Main Methods:

Detailed instructions for expressing fluorescent EMcapsulin reporters in cells or model organisms.
Biochemical quality control measures for assessing reporter expression and integrity.
Guidance on adapting the protocol for custom EMcapsulin construct development.

Main Results:

EMcapsulins manifest as distinct, identifiable shapes in EM images.
Successful labeling of cells and proteins using fluorescent EMcapsulins is demonstrated.
The protocol is adaptable for various research needs and levels of user experience.

Conclusions:

Genetically encoded EMcapsulins provide a versatile and accessible tool for correlative light and electron microscopy.
This protocol empowers researchers to integrate advanced EM techniques into their studies.
The modular nature of EMcapsulins allows for tailored applications in cell biology and beyond.