The recombined cccDNA produced using minicircle technology mimicked HBV genome in structure and function closely

HBV covalently closed circular DNA (cccDNA) is drug-resistant and responsible for viral persistence. To facilitate the development of anti-cccDNA drugs, we developed a minicircle DNA vector (MC)-based technology to produce large quantity of recombined cccDNA (rcccDNA) resembling closely to its wild-...

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Published in:Scientific reports 2016-05, Vol.6 (1), p.25552-25552, Article 25552
Main Authors: Guo, Xiaoyan, Chen, Ping, Hou, Xiaohu, Xu, Wenjuan, Wang, Dan, Wang, Tian-Yan, Zhang, Liping, Zheng, Gang, Gao, Zhi-Liang, He, Cheng-Yi, Zhou, Boping, Chen, Zhi-Ying
Format: Article
Language:English
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Summary:HBV covalently closed circular DNA (cccDNA) is drug-resistant and responsible for viral persistence. To facilitate the development of anti-cccDNA drugs, we developed a minicircle DNA vector (MC)-based technology to produce large quantity of recombined cccDNA (rcccDNA) resembling closely to its wild-type counterpart both in structure and function. The rcccDNA differed to the wild-type cccDNA (wtcccDNA) only in that it carried an extra 36-bp DNA recombinant product attR upstream of the preC/C gene. Using a procedure similar to standard plasmid production, milligrams of rcccDNA can be generated in common laboratories conveniently. The rcccDNA demonstrated many essential biological features of wtcccDNA, including: (1) undergoing nucleation upon nucleus entry; (2) serving as template for production of all HBV RNAs and proteins; (3) deriving virions capable of infecting tree shrew, and subsequently producing viral mRNAs, proteins, rcccDNA and infectious virions. As an example to develop anti-cccDNA drugs, we used the Crispr/Cas9 system to provide clear-cut evidence that rcccDNA was cleaved by this DNA editing tool in vitro. In summary, we have developed a convenient technology to produce large quantity of rcccDNA as a surrogate of wtcccDNA for investigating HBV biology and developing treatment to eradicate this most wide-spreading virus.
ISSN:2045-2322
2045-2322
DOI:10.1038/srep25552