DNA Engineering and Hepatitis B Virus Replication

Recombinant DNA technology is a vital method in human hepatitis B virus (HBV), producing reporter viruses or vectors for gene transferring. Researchers have engineered several genes into the HBV genome for different purposes; however, a systematic analysis of recombinant strategy is lacking. Here, u...

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Autores principales:	Chun-yang Gan, Jing Cui, Wen-lu Zhang, Yu-wei Wang, Ai-long Huang, Jie-li Hu
Formato:	article
Lenguaje:	EN
Publicado:	Frontiers Media S.A. 2021
Materias:	hepatitis B virus recombination replication RNA splicing DNA sequence optimization Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/9a7b7015e35447debdfe5c9e13b9fd78
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spelling	oai:doaj.org-article:9a7b7015e35447debdfe5c9e13b9fd782021-11-11T10:21:51ZDNA Engineering and Hepatitis B Virus Replication1664-302X10.3389/fmicb.2021.783040https://doaj.org/article/9a7b7015e35447debdfe5c9e13b9fd782021-11-01T00:00:00Zhttps://www.frontiersin.org/articles/10.3389/fmicb.2021.783040/fullhttps://doaj.org/toc/1664-302XRecombinant DNA technology is a vital method in human hepatitis B virus (HBV), producing reporter viruses or vectors for gene transferring. Researchers have engineered several genes into the HBV genome for different purposes; however, a systematic analysis of recombinant strategy is lacking. Here, using a 500-bp deletion strategy, we scanned the HBV genome and identified two regions, region I (from nt 2,118 to 2,814) and region II (from nt 99 to 1,198), suitable for engineering. Ten exogenous genes, including puromycin N-acetyl transferase gene (Pac), blasticidin S deaminase gene (BSD), Neomycin-resistance gene (Neo), Gaussia luciferase (Gluc), NanoLuc (Nluc), copGFP, mCherry, UnaG, eGFP, and tTA1, were inserted into these two regions and fused into the open reading frames of hepatitis B core protein (HBC) and hepatitis B surface protein (HBS) via T2A peptide. Recombination of 9 of the 10 genes at region 99–1198 and 5 of the 10 genes at region 2118–2814 supported the formation of relaxed circular (RC) DNA. HBV DNA and HBV RNA assays implied that exogenous genes potentially abrogate RC DNA by inducing the formation of adverse secondary structures. This hypothesis was supported because sequence optimization of the UnaG gene based on HBC sequence rescued RC DNA formation. Findings from this study provide an informative basis and a valuable method for further constructing and optimizing recombinant HBV and imply that DNA sequence might be intrinsically a potential source of selective pressure in the evolution of HBV.Chun-yang GanJing CuiWen-lu ZhangYu-wei WangAi-long HuangJie-li HuFrontiers Media S.A.articlehepatitis B virusrecombinationreplicationRNA splicingDNA sequence optimizationMicrobiologyQR1-502ENFrontiers in Microbiology, Vol 12 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	hepatitis B virus recombination replication RNA splicing DNA sequence optimization Microbiology QR1-502
spellingShingle	hepatitis B virus recombination replication RNA splicing DNA sequence optimization Microbiology QR1-502 Chun-yang Gan Jing Cui Wen-lu Zhang Yu-wei Wang Ai-long Huang Jie-li Hu DNA Engineering and Hepatitis B Virus Replication
description	Recombinant DNA technology is a vital method in human hepatitis B virus (HBV), producing reporter viruses or vectors for gene transferring. Researchers have engineered several genes into the HBV genome for different purposes; however, a systematic analysis of recombinant strategy is lacking. Here, using a 500-bp deletion strategy, we scanned the HBV genome and identified two regions, region I (from nt 2,118 to 2,814) and region II (from nt 99 to 1,198), suitable for engineering. Ten exogenous genes, including puromycin N-acetyl transferase gene (Pac), blasticidin S deaminase gene (BSD), Neomycin-resistance gene (Neo), Gaussia luciferase (Gluc), NanoLuc (Nluc), copGFP, mCherry, UnaG, eGFP, and tTA1, were inserted into these two regions and fused into the open reading frames of hepatitis B core protein (HBC) and hepatitis B surface protein (HBS) via T2A peptide. Recombination of 9 of the 10 genes at region 99–1198 and 5 of the 10 genes at region 2118–2814 supported the formation of relaxed circular (RC) DNA. HBV DNA and HBV RNA assays implied that exogenous genes potentially abrogate RC DNA by inducing the formation of adverse secondary structures. This hypothesis was supported because sequence optimization of the UnaG gene based on HBC sequence rescued RC DNA formation. Findings from this study provide an informative basis and a valuable method for further constructing and optimizing recombinant HBV and imply that DNA sequence might be intrinsically a potential source of selective pressure in the evolution of HBV.
format	article
author	Chun-yang Gan Jing Cui Wen-lu Zhang Yu-wei Wang Ai-long Huang Jie-li Hu
author_facet	Chun-yang Gan Jing Cui Wen-lu Zhang Yu-wei Wang Ai-long Huang Jie-li Hu
author_sort	Chun-yang Gan
title	DNA Engineering and Hepatitis B Virus Replication
title_short	DNA Engineering and Hepatitis B Virus Replication
title_full	DNA Engineering and Hepatitis B Virus Replication
title_fullStr	DNA Engineering and Hepatitis B Virus Replication
title_full_unstemmed	DNA Engineering and Hepatitis B Virus Replication
title_sort	dna engineering and hepatitis b virus replication
publisher	Frontiers Media S.A.
publishDate	2021
url	https://doaj.org/article/9a7b7015e35447debdfe5c9e13b9fd78
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DNA Engineering and Hepatitis B Virus Replication

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