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A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis

Circular single-stranded DNA (ssDNA) viruses are widely distributed globally, infecting diverse hosts ranging from bacteria, archaea, and eukaryotes. Among these, the genome of Banana bunchy top virus (BBTV) comprises at least six circular, ssDNA components that are ∼1 kb in length. Its genome is us...

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Autores principales: Yu, Nai-tong, Zhang, Yu-liang, Wang, Jian-hua, Liu, Zhi-xin
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7396673/
https://www.ncbi.nlm.nih.gov/pubmed/32850712
http://dx.doi.org/10.3389/fbioe.2020.00832
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author Yu, Nai-tong
Zhang, Yu-liang
Wang, Jian-hua
Liu, Zhi-xin
author_facet Yu, Nai-tong
Zhang, Yu-liang
Wang, Jian-hua
Liu, Zhi-xin
author_sort Yu, Nai-tong
collection PubMed
description Circular single-stranded DNA (ssDNA) viruses are widely distributed globally, infecting diverse hosts ranging from bacteria, archaea, and eukaryotes. Among these, the genome of Banana bunchy top virus (BBTV) comprises at least six circular, ssDNA components that are ∼1 kb in length. Its genome is usually amplified and obtained at the DNA level. However, RNA-based techniques to obtain the genome sequence of such multi-component viruses have not been reported. In this study, transcriptome sequencing analysis showed that the full-length of BBTV each genomic component was transcribed into viral mRNA (vmRNA). Accordingly, the near-complete genome of BBTV B2 isolate was assembled using transcriptome sequencing data from virus-infected banana leaves. Assembly analysis of BBTV-derived reads indicated that the full-length sequences were obtained for DNA-R, DNA-U3, DNA-S, DNA-M, DNA-N, NewS2, and Sat4 components, while two gaps (73 and 25 nt) missing in the DNA-C component which was further filled by reverse transcription-PCR (RT-PCR). The RT-qPCR analysis indicated that the vmRNA levels of coding regions were 3.19–103.53 folds higher than those of non-coding regions, implying that the integrity of genome assembly depended on the transcription level of non-coding region. In conclusion, this study proposes a new approach to obtain the genome of nanovirids, and provides insights for studying the transcriptional mechanism of the family Nanoviridae, Genomoviridae, and Geminiviridae.
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spelling pubmed-73966732020-08-25 A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis Yu, Nai-tong Zhang, Yu-liang Wang, Jian-hua Liu, Zhi-xin Front Bioeng Biotechnol Bioengineering and Biotechnology Circular single-stranded DNA (ssDNA) viruses are widely distributed globally, infecting diverse hosts ranging from bacteria, archaea, and eukaryotes. Among these, the genome of Banana bunchy top virus (BBTV) comprises at least six circular, ssDNA components that are ∼1 kb in length. Its genome is usually amplified and obtained at the DNA level. However, RNA-based techniques to obtain the genome sequence of such multi-component viruses have not been reported. In this study, transcriptome sequencing analysis showed that the full-length of BBTV each genomic component was transcribed into viral mRNA (vmRNA). Accordingly, the near-complete genome of BBTV B2 isolate was assembled using transcriptome sequencing data from virus-infected banana leaves. Assembly analysis of BBTV-derived reads indicated that the full-length sequences were obtained for DNA-R, DNA-U3, DNA-S, DNA-M, DNA-N, NewS2, and Sat4 components, while two gaps (73 and 25 nt) missing in the DNA-C component which was further filled by reverse transcription-PCR (RT-PCR). The RT-qPCR analysis indicated that the vmRNA levels of coding regions were 3.19–103.53 folds higher than those of non-coding regions, implying that the integrity of genome assembly depended on the transcription level of non-coding region. In conclusion, this study proposes a new approach to obtain the genome of nanovirids, and provides insights for studying the transcriptional mechanism of the family Nanoviridae, Genomoviridae, and Geminiviridae. Frontiers Media S.A. 2020-07-21 /pmc/articles/PMC7396673/ /pubmed/32850712 http://dx.doi.org/10.3389/fbioe.2020.00832 Text en Copyright © 2020 Yu, Zhang, Wang and Liu. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Bioengineering and Biotechnology
Yu, Nai-tong
Zhang, Yu-liang
Wang, Jian-hua
Liu, Zhi-xin
A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis
title A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis
title_full A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis
title_fullStr A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis
title_full_unstemmed A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis
title_short A New Method to Obtain the Complete Genome Sequence of Multiple-Component Circular ssDNA Viruses by Transcriptome Analysis
title_sort new method to obtain the complete genome sequence of multiple-component circular ssdna viruses by transcriptome analysis
topic Bioengineering and Biotechnology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7396673/
https://www.ncbi.nlm.nih.gov/pubmed/32850712
http://dx.doi.org/10.3389/fbioe.2020.00832
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