Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Related Concept Videos

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...

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by

Same author

Single-cell RNA sequencing of terminal ileal biopsies identifies signatures of Crohn's disease pathogenesis.

Nature genetics·2026

Same author

An Alzheimer's disease-associated common regulatory variant in a <i>PTK2B</i> intron alters microglial function.

iScience·2026

Same author

Optimisation of Xenium automated in situ sequencing for PAXgene-fixed tissue samples.

Journal of histotechnology·2026

Same author

The Human Pleiotropic Map of GWAS Associations and Therapeutic Implications.

bioRxiv : the preprint server for biology·2026

Same author

Epigenetic biomarkers in neurodegenerative diseases: from molecular signatures to therapeutic targets.

Trends in neurosciences·2026

Same author

Genome-scale mapping of variant, enhancer and gene function in primary human CD4+ T cells.

bioRxiv : the preprint server for biology·2026

Same journal

Cap 2'-O-methyltransferase CMTR2 regulates male meiosis independent of its methyltransferase activity.

Nucleic acids research·2026

Same journal

APE1 binds and processes abasic sites present in i-motif DNA and cooperates with PCBP1 in maintenance of telomeric stability.

Nucleic acids research·2026

Same journal

Acquisition of a novel restriction modification system regulates genetic flux and gene expression in the hypervirulent and globally disseminated CC17 lineage of group B Streptococcus.

Nucleic acids research·2026

Same journal

Trans-species microRNAs from the parasitic plant Cuscuta campestris specifically avoid loading onto self Argonautes.

Nucleic acids research·2026

Same journal

Neurochondrin promotes U5 snRNP maturation by regulating AAR2 release from PRPF8.

Nucleic acids research·2026

Same journal

Elongationless start-stop elements are stress-resilient translation gates that are more repressive than uTranslons.

Nucleic acids research·2026

See all related articles

Search research articles

Related Experiment Video

Updated: Sep 11, 2025

Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites

Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites

Published on: March 22, 2016

ONE-STEP tagging: a versatile method for rapid site-specific integration by simultaneous reagent delivery.

Valentina Migliori¹, Michaela B Bruntraeger¹, Ivan S Gyulev¹

¹Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, CB10 1SA, United Kingdom.

Nucleic Acids Research

|August 18, 2025

Summary

This summary is machine-generated.

We developed ONE-STEP tagging, a novel genome editing tool combining CRISPR-Cas9 and Bxb1 integrase for precise protein tagging. This method efficiently tags proteins in various cells, including stem and T cells, advancing genome engineering for research and therapy.

More Related Videos

One-step CRISPR-based Strategy for Endogenous Gene Tagging in Drosophila melanogaster

One-step CRISPR-based Strategy for Endogenous Gene Tagging in Drosophila melanogaster

Published on: January 26, 2024

Inducible LAP-tagged Stable Cell Lines for Investigating Protein Function, Spatiotemporal Localization and Protein Interaction Networks

Inducible LAP-tagged Stable Cell Lines for Investigating Protein Function, Spatiotemporal Localization and Protein Interaction Networks

Published on: December 24, 2016

Related Experiment Videos

Last Updated: Sep 11, 2025

Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites

Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites

Published on: March 22, 2016

One-step CRISPR-based Strategy for Endogenous Gene Tagging in Drosophila melanogaster

One-step CRISPR-based Strategy for Endogenous Gene Tagging in Drosophila melanogaster

Published on: January 26, 2024

Inducible LAP-tagged Stable Cell Lines for Investigating Protein Function, Spatiotemporal Localization and Protein Interaction Networks

Inducible LAP-tagged Stable Cell Lines for Investigating Protein Function, Spatiotemporal Localization and Protein Interaction Networks

Published on: December 24, 2016

Area of Science:

* Molecular Biology
* Genome Engineering
* Gene Editing

Background:

* Precise protein tagging is crucial for understanding cellular functions and developing therapeutics.
* Existing genome editing methods face challenges in efficiency, scalability, and cargo capacity.
* Site-specific integration offers advantages over random insertion for stable gene expression.

Purpose of the Study:

* To introduce and validate a novel, versatile genome editing method called ONE-STEP tagging.
* To demonstrate the efficiency, precision, and scalability of ONE-STEP tagging for protein labeling.
* To showcase the applicability of ONE-STEP tagging in various cell types, including primary T cells, for therapeutic applications.

Main Methods:

* Combined CRISPR-Cas9 for targeting with Bxb1 integrase for site-specific DNA integration.
* Optimized Bxb1 integrase and used a DNA-PK inhibitor to enhance integration efficiency.
* Applied the method in human-induced pluripotent stem cells (hiPSCs), cancer cells, and primary T cells.
* Tested integration of large DNA constructs (up to 14.4 kb) and multiplexed tagging.
* Demonstrated simultaneous gene editing (e.g., knockout) at secondary loci.

Main Results:

* Achieved up to ~90% integration efficiency at the ACTR10 locus in hiPSCs.
* Demonstrated robust performance across different cell types and genomic loci.
* Successfully integrated a large 14.4 kb DNA construct with 16.6% efficiency.
* Enabled multiplexed protein tagging and simultaneous secondary gene editing.
* Applied ONE-STEP tagging in primary T cells for CAR T-cell therapy development, including B2M knockout.

Conclusions:

* ONE-STEP tagging is a highly efficient, precise, and scalable method for endogenous protein tagging.
* The method is versatile, supporting large DNA cargo and multiplexing capabilities.
* ONE-STEP tagging shows significant potential for both basic research and therapeutic genome engineering, particularly in cell-based therapies.