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

In vitro Mutagenesis

In vitro Mutagenesis

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.

PCR

PCR

Real Time RT-PCR

Real Time RT-PCR

Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...

In-vitro Mutagenesis

In-vitro Mutagenesis

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.

You might also read

Related Articles

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

Sort by

Same author

Structural basis for transport and inhibition of nucleotide sugar transport in pathogenic fungi.

Nature communications·2026

Same author

Structural basis for pH-responsive amino acid transport via SLC7A4.

Nature communications·2026

Same author

Structural basis for prostaglandin and drug transport via SLCO2A1.

Nature communications·2026

Same author

Mechanism of phosphoinositide regulation of lysosomal pH via inhibition of CLC-7.

bioRxiv : the preprint server for biology·2025

Same author

Rad53 regulates RNase H1, which promotes DNA replication through sites of transcription-replication conflict.

Cell reports·2025

Same author

The INO80 chromatin remodeller facilitates DNA damage bypass via postreplicative gap repair.

The EMBO journal·2025

Same journal

Tracking Synthetic Adhesins on Bacterial Surfaces with Immunofluorescence Microscopy.

Methods in molecular biology (Clifton, N.J.)·2026

Same journal

Post-Selection Methods for Analyzing mRNA Display Selections and Optimization of Hits.

Methods in molecular biology (Clifton, N.J.)·2026

Same journal

High-Performance Computing in Tandem Mass Spectrometry (MS/MS) Peptide Identification.

Methods in molecular biology (Clifton, N.J.)·2026

Same journal

Engineering and Adapting Disulfide-Containing Proteins to Enable Intracellular Functionality.

Methods in molecular biology (Clifton, N.J.)·2026

Same journal

AI-Driven Protein Research: From Prediction to Design.

Methods in molecular biology (Clifton, N.J.)·2026

Same journal

Methods for the In Vitro Selection of Protein and Peptide Libraries Using mRNA Display.

Methods in molecular biology (Clifton, N.J.)·2026

See all related articles

Search research articles

Related Experiment Video

Updated: May 19, 2026

Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control

Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control

Published on: March 30, 2015

In vitro PCNA modification assays.

Joanne L Parker¹, Helle D Ulrich

¹Clare Hall Laboratories, Cancer Research UK London Research Institute, South Mimms, UK.

Methods in Molecular Biology (Clifton, N.J.)

|September 4, 2012

Summary

This summary is machine-generated.

Researchers purified enzymes and proteins to study how modifications like ubiquitylation and SUMOylation of proliferating cell nuclear antigen (PCNA) help repair DNA damage during replication.

More Related Videos

Digital Polymerase Chain Reaction Assay for the Genetic Variation in a Sporadic Familial Adenomatous Polyposis Patient Using the Chip-in-a-tube Format

Digital Polymerase Chain Reaction Assay for the Genetic Variation in a Sporadic Familial Adenomatous Polyposis Patient Using the Chip-in-a-tube Format

Published on: August 20, 2018

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

Published on: November 3, 2010

Related Experiment Videos

Last Updated: May 19, 2026

Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control

Development of a Quantitative Recombinase Polymerase Amplification Assay with an Internal Positive Control

Published on: March 30, 2015

Digital Polymerase Chain Reaction Assay for the Genetic Variation in a Sporadic Familial Adenomatous Polyposis Patient Using the Chip-in-a-tube Format

Digital Polymerase Chain Reaction Assay for the Genetic Variation in a Sporadic Familial Adenomatous Polyposis Patient Using the Chip-in-a-tube Format

Published on: August 20, 2018

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

Published on: November 3, 2010

Area of Science:

Molecular Biology
Biochemistry
Genetics

Background:

The replicative sliding clamp, proliferating cell nuclear antigen (PCNA), is crucial for DNA replication and repair.
Post-translational modifications of PCNA, including ubiquitylation and SUMOylation, play significant roles in processing DNA damage during replication.
Understanding the enzymes and mechanisms governing these modifications is essential for comprehending DNA damage tolerance pathways.

Purpose of the Study:

To investigate the properties, interactions, substrate recognition, and regulatory mechanisms of PCNA conjugation enzymes.
To establish in vitro reconstitution systems for studying ubiquitylation and SUMOylation of PCNA.
To develop protocols for purifying modified PCNA for further biochemical analysis.

Main Methods:

Purification of relevant enzymes and accessory proteins from E. coli and S. cerevisiae.
Setting up small-scale in vitro ubiquitylation and SUMOylation reactions using purified budding yeast PCNA.
Development of a method for purifying monoubiquitylated PCNA.

Main Results:

Successful purification of key enzymes and accessory proteins involved in PCNA modification.
Establishment of functional in vitro systems for reconstituting ubiquitylation and SUMOylation of PCNA.
A protocol for obtaining monoubiquitylated PCNA suitable for subsequent biochemical investigations.

Conclusions:

The described methods enable detailed biochemical studies of PCNA modification pathways.
These reconstituted systems facilitate the investigation of enzyme properties, substrate interactions, and regulatory mechanisms.
The purification of monoubiquitylated PCNA provides a valuable resource for further research into DNA damage processing.