Table 1.

Detection methods for HBV DNA integration.

Technique	Suitable Uses	Advantages	Limitations	References
Southern blot hybridization	HBV DNA integration detection in liver samples with highly expanded hepatocyte clones	Low cost	➢Time consuming ➢No sequence information ➢Dependent on restriction enzyme sites ➢Low sensitivity	[47,48,49,50]
Direct cloning and Sanger sequencing	Defining structure of HBV integration in liver samples with highly expanded hepatocyte clones	Definition of the sequences of integrated HBV DNA and the adjacent cellular DNA	➢Not suitable for screening a large number of unknown virus–cell junctions ➢Low throughput ➢Dependent on restriction enzyme sites for cloning	[50,51]
Alu PCR	Detecting and sequencing of HBV integration in clonally expanded hepatocytes	Inexpensive	➢Can only effectively detect HBV integration near Alu sequences ➢Dependent on Alu sequences ➢No integration quantification ➢Biased towards larger clones	[52,53,54,55,56,57]
Inverse PCR	Detecting and quantifying HBV DNA integrations in small hepatocyte clones (single copy virus–cell junctions can be detected)	➢Absolute integration quantification ➢High sensitivity	➢Dependent on restriction enzyme sites for detection of virus–host DNA junction ➢Detection of integrations that occur between nucleotides ~1650 and ~1850 of viral genome	[39,41,58,59,60]
Whole genome sequencing (WGS)	Sensitive and comprehensive inthe identification of viral integrants across the human genome	Full genome coverage	➢Low depth ➢High cost ➢No absolute quantification	[61,62,63]
Whole-exome sequencing	Detection of HBV integration in coding regions	Greater depth than WGS	➢Coverage limited to coding regions ➢No absolute integration quantification	[64,65,66,67]
RNA Sequencing	Sensitive and comprehensive in the identification of viral integrants across the human transcriptome	➢Greater depth than WGS ➢Data on transcriptional activity	➢Coverage limited to expressed coding regions ➢No absolute integration quantification ➢Biased towards more highly expressed genes	[42,65,66,67,68]
Capture-enriched next generation sequencing	High-throughput viral integration detection method	➢Cost-effective compared with WGS ➢Quicker and less laborious than PCR-based methods	➢Lower sensitivity than PCR-based methods ➢Lower specificity than PCR-based methods ➢Shorter fragments are captured with higher specificity than longer ones	[42,69]