Viral metagenomics: Analysis of begomoviruses by illumina high-throughput sequencing

Handle URI:
http://hdl.handle.net/10754/325365
Title:
Viral metagenomics: Analysis of begomoviruses by illumina high-throughput sequencing
Authors:
Idris, Ali; Al-Saleh, Mohammed; Piatek, Marek J.; Al-Shahwan, Ibrahim; Ali, Shahjahan; Brown, Judith K.
Abstract:
Traditional DNA sequencing methods are inefficient, lack the ability to discern the least abundant viral sequences, and ineffective for determining the extent of variability in viral populations. Here, populations of single-stranded DNA plant begomoviral genomes and their associated beta- and alpha-satellite molecules (virus-satellite complexes) (genus, Begomovirus; family, Geminiviridae) were enriched from total nucleic acids isolated from symptomatic, field-infected plants, using rolling circle amplification (RCA). Enriched virus-satellite complexes were subjected to Illumina-Next Generation Sequencing (NGS). CASAVA and SeqMan NGen programs were implemented, respectively, for quality control and for de novo and reference-guided contig assembly of viral-satellite sequences. The authenticity of the begomoviral sequences, and the reproducibility of the Illumina-NGS approach for begomoviral deep sequencing projects, were validated by comparing NGS results with those obtained using traditional molecular cloning and Sanger sequencing of viral components and satellite DNAs, also enriched by RCA or amplified by polymerase chain reaction. As the use of NGS approaches, together with advances in software development, make possible deep sequence coverage at a lower cost; the approach described herein will streamline the exploration of begomovirus diversity and population structure from naturally infected plants, irrespective of viral abundance. This is the first report of the implementation of Illumina-NGS to explore the diversity and identify begomoviral-satellite SNPs directly from plants naturally-infected with begomoviruses under field conditions. 2014 by the authors; licensee MDPI, Basel, Switzerland.
KAUST Department:
Center for Desert Agriculture
Citation:
Idris A, Al-Saleh M, Piatek M, Al-Shahwan I, Ali S, et al. (2014) Viral Metagenomics: Analysis of Begomoviruses by Illumina High-Throughput Sequencing. Viruses 6: 1219-1236. doi:10.3390/v6031219.
Publisher:
MDPI
Journal:
Viruses
Issue Date:
12-Mar-2014
DOI:
10.3390/v6031219
PubMed ID:
24625811
PubMed Central ID:
PMC3970147
Type:
Article
ISSN:
19994915
Appears in Collections:
Articles; Center for Desert Agriculture

Full metadata record

DC FieldValue Language
dc.contributor.authorIdris, Alien
dc.contributor.authorAl-Saleh, Mohammeden
dc.contributor.authorPiatek, Marek J.en
dc.contributor.authorAl-Shahwan, Ibrahimen
dc.contributor.authorAli, Shahjahanen
dc.contributor.authorBrown, Judith K.en
dc.date.accessioned2014-08-27T09:49:19Z-
dc.date.available2014-08-27T09:49:19Z-
dc.date.issued2014-03-12en
dc.identifier.citationIdris A, Al-Saleh M, Piatek M, Al-Shahwan I, Ali S, et al. (2014) Viral Metagenomics: Analysis of Begomoviruses by Illumina High-Throughput Sequencing. Viruses 6: 1219-1236. doi:10.3390/v6031219.en
dc.identifier.issn19994915en
dc.identifier.pmid24625811en
dc.identifier.doi10.3390/v6031219en
dc.identifier.urihttp://hdl.handle.net/10754/325365en
dc.description.abstractTraditional DNA sequencing methods are inefficient, lack the ability to discern the least abundant viral sequences, and ineffective for determining the extent of variability in viral populations. Here, populations of single-stranded DNA plant begomoviral genomes and their associated beta- and alpha-satellite molecules (virus-satellite complexes) (genus, Begomovirus; family, Geminiviridae) were enriched from total nucleic acids isolated from symptomatic, field-infected plants, using rolling circle amplification (RCA). Enriched virus-satellite complexes were subjected to Illumina-Next Generation Sequencing (NGS). CASAVA and SeqMan NGen programs were implemented, respectively, for quality control and for de novo and reference-guided contig assembly of viral-satellite sequences. The authenticity of the begomoviral sequences, and the reproducibility of the Illumina-NGS approach for begomoviral deep sequencing projects, were validated by comparing NGS results with those obtained using traditional molecular cloning and Sanger sequencing of viral components and satellite DNAs, also enriched by RCA or amplified by polymerase chain reaction. As the use of NGS approaches, together with advances in software development, make possible deep sequence coverage at a lower cost; the approach described herein will streamline the exploration of begomovirus diversity and population structure from naturally infected plants, irrespective of viral abundance. This is the first report of the implementation of Illumina-NGS to explore the diversity and identify begomoviral-satellite SNPs directly from plants naturally-infected with begomoviruses under field conditions. 2014 by the authors; licensee MDPI, Basel, Switzerland.en
dc.language.isoenen
dc.publisherMDPIen
dc.rightsThis article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/3.0/).en
dc.subjectGeminivirusen
dc.subjectGenome enrichmenten
dc.subjectIllumina sequencingen
dc.subjectssDNA virusesen
dc.subjectViral genome assemblyen
dc.subjectDNA helicaseen
dc.subjectsatellite DNAen
dc.subjectsingle stranded DNAen
dc.subjectBegomovirusen
dc.subjectcontrolled studyen
dc.subjectDNA sequenceen
dc.subjectgene mappingen
dc.subjectgenetic codeen
dc.subjectgenome analysisen
dc.subjecthigh throughput sequencingen
dc.subjectmetagenomicsen
dc.subjectNicotiana benthamianaen
dc.subjectnucleotide sequenceen
dc.subjectparsimony analysisen
dc.subjectphylogenyen
dc.subjectplant virusen
dc.subjectpolymerase chain reactionen
dc.subjectrestriction fragment length polymorphismen
dc.subjectsingle nucleotide polymorphismen
dc.subjecttomato yellow leaf curl betasatelliteen
dc.subjectTomato yellow leaf curl virusen
dc.subjectvirus genomeen
dc.titleViral metagenomics: Analysis of begomoviruses by illumina high-throughput sequencingen
dc.typeArticleen
dc.contributor.departmentCenter for Desert Agricultureen
dc.identifier.journalVirusesen
dc.identifier.pmcidPMC3970147en
dc.eprint.versionPublisher's Version/PDFen
dc.contributor.institutionDepartment of Plant Protection, King Saud University, Riyadh 11451, Saudi Arabiaen
dc.contributor.institutionSchool of Plant Sciences, The University of Arizona, Tucson, AZ 85721, United Statesen
dc.contributor.affiliationKing Abdullah University of Science and Technology (KAUST)en
kaust.authorIdris, Alien
kaust.authorAli, Shahjahanen
kaust.authorPiatek, Marek J.en

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