Unit-length, single-stranded circular DNAs of both polarity of begomoviruses are generated in Escherichia coli harboring phage M13-cloned begomovirus genome with single copy of replication origin
Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA replication process of begomoviruses in bacteria. However, previous studies indicated that the replication of begomovirus DNAs in prokaryotes requires...
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| Published in | Virus research Vol. 125; no. 1; pp. 14 - 28 |
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| Main Authors | , , , , , |
| Format | Journal Article |
| Language | English |
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Netherlands
Elsevier B.V
01.04.2007
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| Abstract | Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA replication process of begomoviruses in bacteria. However, previous studies indicated that the replication of begomovirus DNAs in prokaryotes requires tandem constructs of viral genomes with at least two copies of the origin of replication (
ori). In this study, phage M13 vector harboring the unit-length genome with only a single copy of
ori of a mono-partite begomovirus,
Ageratum yellow vein virus PD isolate (AYVV-[PD]), was constructed and used to investigate the replication of AYVV-[PD] DNAs in
Escherichia coli. The generation of single-stranded, circular DNAs (sscDNAs) corresponding to the unit-length AYVV-[PD] genome of both polarity was observed and verified. Replication-associated (Rep) protein of AYVV-[PD] was detected only in bacteria generating the corresponding sscDNAs, whereas disruption of the Rep gene abolished the phenomenon. The results suggested that a single copy of
ori is sufficient for the prokaryotes to support the generation of unit-length, genomic sscDNAs of begomoviruses, which requires the presence of functional Rep protein. |
|---|---|
| AbstractList | Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA replication process of begomoviruses in bacteria. However, previous studies indicated that the replication of begomovirus DNAs in prokaryotes requires tandem constructs of viral genomes with at least two copies of the origin of replication (ori). In this study, phage M13 vector harboring the unit-length genome with only a single copy of ori of a mono-partite begomovirus, Ageratum yellow vein virus PD isolate (AYVV-[PD]), was constructed and used to investigate the replication of AYVV-[PD] DNAs in Escherichia coli. The generation of single-stranded, circular DNAs (sscDNAs) corresponding to the unit-length AYVV-[PD] genome of both polarity was observed and verified. Replication-associated (Rep) protein of AYVV-[PD] was detected only in bacteria generating the corresponding sscDNAs, whereas disruption of the Rep gene abolished the phenomenon. The results suggested that a single copy of ori is sufficient for the prokaryotes to support the generation of unit-length, genomic sscDNAs of begomoviruses, which requires the presence of functional Rep protein. Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA replication process of begomoviruses in bacteria. However, previous studies indicated that the replication of begomovirus DNAs in prokaryotes requires tandem constructs of viral genomes with at least two copies of the origin of replication ( ori). In this study, phage M13 vector harboring the unit-length genome with only a single copy of ori of a mono-partite begomovirus, Ageratum yellow vein virus PD isolate (AYVV-[PD]), was constructed and used to investigate the replication of AYVV-[PD] DNAs in Escherichia coli. The generation of single-stranded, circular DNAs (sscDNAs) corresponding to the unit-length AYVV-[PD] genome of both polarity was observed and verified. Replication-associated (Rep) protein of AYVV-[PD] was detected only in bacteria generating the corresponding sscDNAs, whereas disruption of the Rep gene abolished the phenomenon. The results suggested that a single copy of ori is sufficient for the prokaryotes to support the generation of unit-length, genomic sscDNAs of begomoviruses, which requires the presence of functional Rep protein. Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA replication process of begomoviruses in bacteria. However, previous studies indicated that the replication of begomovirus DNAs in prokaryotes requires tandem constructs of viral genomes with at least two copies of the origin of replication (ori). In this study, phage M13 vector harboring the unit-length genome with only a single copy of ori of a mono-partite begomovirus, Ageratum yellow vein virus PD isolate (AYVV-[PD]), was constructed and used to investigate the replication of AYVV-[PD] DNAs in Escherichia coli. The generation of single-stranded, circular DNAs (sscDNAs) corresponding to the unit-length AYVV-[PD] genome of both polarity was observed and verified. Replication-associated (Rep) protein of AYVV-[PD] was detected only in bacteria generating the corresponding sscDNAs, whereas disruption of the Rep gene abolished the phenomenon. The results suggested that a single copy of ori is sufficient for the prokaryotes to support the generation of unit-length, genomic sscDNAs of begomoviruses, which requires the presence of functional Rep protein.Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA replication process of begomoviruses in bacteria. However, previous studies indicated that the replication of begomovirus DNAs in prokaryotes requires tandem constructs of viral genomes with at least two copies of the origin of replication (ori). In this study, phage M13 vector harboring the unit-length genome with only a single copy of ori of a mono-partite begomovirus, Ageratum yellow vein virus PD isolate (AYVV-[PD]), was constructed and used to investigate the replication of AYVV-[PD] DNAs in Escherichia coli. The generation of single-stranded, circular DNAs (sscDNAs) corresponding to the unit-length AYVV-[PD] genome of both polarity was observed and verified. Replication-associated (Rep) protein of AYVV-[PD] was detected only in bacteria generating the corresponding sscDNAs, whereas disruption of the Rep gene abolished the phenomenon. The results suggested that a single copy of ori is sufficient for the prokaryotes to support the generation of unit-length, genomic sscDNAs of begomoviruses, which requires the presence of functional Rep protein. |
| Author | Lai, Yi-Chin Yang, Su-Hua Hsu, Yau-Heiu Lin, Na-Sheng Hu, Chung-Chi Wu, Chia-Ying |
| Author_xml | – sequence: 1 givenname: Chia-Ying surname: Wu fullname: Wu, Chia-Ying organization: Graduate Institute of Biotechnology, National Chung Hsing University, 250 Kuo-Kuang Road, Taichung-402, Taiwan – sequence: 2 givenname: Su-Hua surname: Yang fullname: Yang, Su-Hua organization: Graduate Institute of Biotechnology, National Chung Hsing University, 250 Kuo-Kuang Road, Taichung-402, Taiwan – sequence: 3 givenname: Yi-Chin surname: Lai fullname: Lai, Yi-Chin organization: Graduate Institute of Biotechnology, National Chung Hsing University, 250 Kuo-Kuang Road, Taichung-402, Taiwan – sequence: 4 givenname: Na-Sheng surname: Lin fullname: Lin, Na-Sheng organization: Institute of Plant and Microbial Biology, Academia Sinica, Nankang, Taipei 115, Taiwan – sequence: 5 givenname: Yau-Heiu surname: Hsu fullname: Hsu, Yau-Heiu organization: Graduate Institute of Biotechnology, National Chung Hsing University, 250 Kuo-Kuang Road, Taichung-402, Taiwan – sequence: 6 givenname: Chung-Chi surname: Hu fullname: Hu, Chung-Chi email: cchu@dragon.nchu.edu.tw organization: Graduate Institute of Biotechnology, National Chung Hsing University, 250 Kuo-Kuang Road, Taichung-402, Taiwan |
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| Keywords | Phage M13 Replication-associated protein Single-stranded circular DNA Escherichia coli Begomoviruses |
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| Snippet | Replication of genomic DNAs of plant-pathogenic begomoviruses has been demonstrated in prokaryotes, which supported the possibility of analyzing DNA... |
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| SubjectTerms | Ageratum yellow vein virus Bacteriophage M13 - genetics Begomovirus Begomovirus - genetics Begomoviruses Coliphages - genetics DNA, Circular - genetics DNA, Circular - metabolism DNA, Single-Stranded DNA, Viral - genetics DNA, Viral - metabolism Escherichia coli Escherichia coli - genetics Escherichia coli - virology Genetic Engineering - methods Genome, Viral Phage M13 Plasmids - genetics Replication Origin Replication-associated protein Single-stranded circular DNA |
| Title | Unit-length, single-stranded circular DNAs of both polarity of begomoviruses are generated in Escherichia coli harboring phage M13-cloned begomovirus genome with single copy of replication origin |
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