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Related Experiment Video

Updated: Jul 9, 2025

Detection of Low Copy Number Integrated Viral DNA Formed by In Vitro Hepatitis B Infection
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Detection of Low Copy Number Integrated Viral DNA Formed by In Vitro Hepatitis B Infection

Published on: November 7, 2018

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A versatile method to profile hepatitis B virus DNA integration.

Kento Fukano1,2, Kousho Wakae1, Naganori Nao3,4,5

  • 1Department of Virology II, National Institute of Infectious Diseases, Tokyo, Japan.

Hepatology Communications
|December 5, 2023
PubMed
Summary

A new method called RAISING efficiently detects Hepatitis B Virus (HBV) DNA integration sites in host genomes. This cost-effective technique aids in understanding HBV-associated liver cancer and diagnosis.

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Last Updated: Jul 9, 2025

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Area of Science:

  • Molecular Biology
  • Genomics
  • Hepatology

Background:

  • Hepatocellular carcinoma (HCC) is often linked to Hepatitis B Virus (HBV) DNA integration into the host genome, a key factor in hepatocarcinogenesis.
  • Existing methods for detecting HBV integration sites have limitations in sensitivity, cost, and throughput.
  • There is a need for sensitive, comprehensive, and cost-effective methods to detect viral integration sites.

Purpose of the Study:

  • To investigate the efficiency of RAISING (Rapid Amplification of Integration Site without Interference by Genomic DNA contamination) for detecting HBV DNA integration.
  • To establish RAISING as a simple, inexpensive, and highly sensitive method for identifying viral integration.

Main Methods:

  • RAISING utilizes virus-specific primers to amplify HBV-integrated fragments, covering the entire HBV genome.
  • Products were sequenced using Illumina, and HBV-human junction sequences were identified in HCC cell lines.
  • Validation was performed using junction-specific nested PCR and droplet digital PCR in various models and clinical specimens.

Main Results:

  • RAISING-sequencing successfully identified HBV-human junction sequences and their frequencies in HCC cell lines.
  • The method demonstrated superior performance in concentrating junction sequences compared to hybrid capture-sequencing.
  • RAISING-sequencing accurately determined de novo integration sites in various cell types, animal models, and clinical samples.

Conclusions:

  • RAISING provides a cost-effective and sensitive approach for profiling HBV-human junction sequences, requiring less sequencing data.
  • This method is expected to advance research in basic HBV integration mechanisms.
  • RAISING is poised to become a valuable tool for clinical diagnosis in HBV-related diseases.