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Related Experiment Video

Updated: Jul 13, 2026

Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

Published on: May 2, 2025

Molecular and structural transactions at human DNA replication origins.

Arturo Falaschi¹, Gulnara Abdurashidova, Oscar Sandoval

¹International Centre for Genetic Engineering and Biotechnology, Trieste, Italy. falaschi@icgeb.org

Cell Cycle (Georgetown, Tex.)

|July 12, 2007

Summary

Metazoan DNA replication origins lack sequence specificity, unlike yeast. Topoisomerases I and II play crucial roles in origin definition and cell-cycle-dependent activation by interacting with the origin DNA.

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Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

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G2-seq: A High Throughput Sequencing-based Technique for Identifying Late Replicating Regions of the Genome

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Published on: March 22, 2018

Area of Science:

Molecular Biology
Genetics
Cell Biology

Background:

Metazoan DNA replication origins are sites of complex protein-DNA interactions.
The Origin Recognition Complex (ORC) is essential but lacks sequence specificity in metazoans, unlike in yeast.
Metazoan genomes lack identifiable origin consensus sequences.

Purpose of the Study:

To investigate the role of DNA topology and topoisomerases in metazoan DNA replication origin function.
To understand the cell-cycle-dependent interactions of topoisomerases with replication origins.

Main Methods:

Analysis of protein-DNA interactions at the human lamin B2 origin.
In vitro binding assays for topoisomerase I and II.
Cell-cycle analysis of enzyme presence at the origin.

Main Results:

Topoisomerases I and II interact specifically with the human lamin B2 origin in a cell-cycle-dependent manner.
These topoisomerases compete with ORC2 for binding sites at the origin.
Topoisomerase II facilitates prereplicative complex assembly, while topoisomerase I enables initiation of DNA synthesis.

Conclusions:

Topoisomerases are critical for defining metazoan replication origins and regulating their activation.
Their cell-cycle-modulated interactions with origin DNA are essential for precise replication control.