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| Next Generation Transcriptome Sequencing and Quantitative Real-Time PCR Technologies for Characterisation of the Bemisia tabaci Asia 1 mtCOI Phylogenetic Clade |
| Susan Seal, Mitulkumar V Patel, Carl Collins, John Colvin , David Bailey |
| 1.Natural Resources Institute, University of Greenwich, Chatham Maritime Kent ME4 4TB, United Kingdom |
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摘要 A programme of functional genomics research is underway at the University of Greenwich, UK, to develop and apply genomics technologies to characterise an economically-important but under-researched Bemisia tabaci (Hemiptera: Aleyrodidae), the Asia 1 mtCOI phylogenetic group. A comparison of this putative species from India with other important B. tabaci populations and insect species may provide targets for the development of more effective whitefly control strategies. As a first step, next-generation sequencing (NGS) has been used to survey the transcriptome of adult female whitefly, with high quality RNA preparations being used to generate cDNA libraries for NGS using the Roche 454 Titanium DNA sequencing platform. Contig assemblies constructed from the resultant sequences (301 094 reads) using the software program CLC Genomics Workbench generated 3 821 core contigs. Comparison of a selection of these contigs with related sequences from other B. tabaci genetic groups has revealed good alignment for some genes (e.g., HSP90) but misassemblies in other datasets (e.g., the vitellogenin gene family), highlighting the need for manual curation as well as collaborative international efforts to obtain accurate assemblies from the existing next generation sequence datasets. Nevertheless, data emerging from the NGS has facilitated the development of accurate and reliable methods for analysing gene expression based on quantitative real-time RT-PCR, illustrating the power of this approach to enable rapid expression analyses in an organism for which a complete genome sequence is currently lacking.
Abstract A programme of functional genomics research is underway at the University of Greenwich, UK, to develop and apply genomics technologies to characterise an economically-important but under-researched Bemisia tabaci (Hemiptera: Aleyrodidae), the Asia 1 mtCOI phylogenetic group. A comparison of this putative species from India with other important B. tabaci populations and insect species may provide targets for the development of more effective whitefly control strategies. As a first step, next-generation sequencing (NGS) has been used to survey the transcriptome of adult female whitefly, with high quality RNA preparations being used to generate cDNA libraries for NGS using the Roche 454 Titanium DNA sequencing platform. Contig assemblies constructed from the resultant sequences (301 094 reads) using the software program CLC Genomics Workbench generated 3 821 core contigs. Comparison of a selection of these contigs with related sequences from other B. tabaci genetic groups has revealed good alignment for some genes (e.g., HSP90) but misassemblies in other datasets (e.g., the vitellogenin gene family), highlighting the need for manual curation as well as collaborative international efforts to obtain accurate assemblies from the existing next generation sequence datasets. Nevertheless, data emerging from the NGS has facilitated the development of accurate and reliable methods for analysing gene expression based on quantitative real-time RT-PCR, illustrating the power of this approach to enable rapid expression analyses in an organism for which a complete genome sequence is currently lacking.
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Received: 15 April 2011
Accepted:
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| Fund: Funding for the studies described was provided by the University of Greenwich Proof of Concept and Research Funds, UK (E0162/RAE-NRI-009/09 and K0070). |
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Corresponding Authors:
Correspondence Susan Seal, Tel: +44-1634-883602, Fax: +44-1634-883379, E-mail: s.e.seal@gre.ac.uk
E-mail: s.e.seal@gre.ac.uk
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| About author: Susan Seal, Tel: +44-1634-883602, Fax: +44-1634-883379, E-mail: s.e.seal@gre.ac.uk |
Cite this article:
Susan Seal, Mitulkumar V Patel, Carl Collins, John Colvin , David Bailey.
2012.
Next Generation Transcriptome Sequencing and Quantitative Real-Time PCR Technologies for Characterisation of the Bemisia tabaci Asia 1 mtCOI Phylogenetic Clade. Journal of Integrative Agriculture, 11(2): 281-292.
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