A targeted enrichment strategy for massively parallel sequencing of angiosperm plastid genomes1
Identifieur interne : 000F70 ( Main/Exploration ); précédent : 000F69; suivant : 000F71A targeted enrichment strategy for massively parallel sequencing of angiosperm plastid genomes1
Auteurs : Gregory W. Stull [États-Unis] ; Michael J. Moore [États-Unis] ; Venkata S. Mandala [États-Unis] ; Norman A. Douglas [États-Unis] ; Heather-Rose Kates [États-Unis] ; Xinshuai Qi [République populaire de Chine] ; Samuel F. Brockington [Royaume-Uni] ; Pamela S. Soltis [États-Unis] ; Douglas E. Soltis [États-Unis] ; Matthew A. Gitzendanner [États-Unis]Source :
- Applications in Plant Sciences [ 2168-0450 ] ; 2013.
Abstract
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•
•
Url:
DOI: 10.3732/apps.1200497
PubMed: 25202518
PubMed Central: 4105372
Affiliations:
- Royaume-Uni, République populaire de Chine, États-Unis
- Angleterre, Angleterre de l'Est, Floride, Ohio, Zhejiang
- Cambridge, Hangzhou
- Université de Cambridge, Université de Zhejiang
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Le document en format XML
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<front><div type="abstract" xml:lang="en"><p>• <italic>Premise of the study:</italic>
We explored a targeted enrichment strategy to facilitate rapid and low-cost next-generation sequencing (NGS) of numerous complete plastid genomes from across the phylogenetic breadth of angiosperms.</p>
<p>• <italic>Methods and Results:</italic>
A custom RNA probe set including the complete sequences of 22 previously sequenced eudicot plastomes was designed to facilitate hybridization-based targeted enrichment of eudicot plastid genomes. Using this probe set and an Agilent SureSelect targeted enrichment kit, we conducted an enrichment experiment including 24 angiosperms (22 eudicots, two monocots), which were subsequently sequenced on a single lane of the Illumina GAIIx with single-end, 100-bp reads. This approach yielded nearly complete to complete plastid genomes with exceptionally high coverage (mean coverage: 717×), even for the two monocots.</p>
<p>• <italic>Conclusions:</italic>
Our enrichment experiment was highly successful even though many aspects of the capture process employed were suboptimal. Hence, significant improvements to this methodology are feasible. With this general approach and probe set, it should be possible to sequence more than 300 essentially complete plastid genomes in a single Illumina GAIIx lane (achieving ∼50× mean coverage). However, given the complications of pooling numerous samples for multiplex sequencing and the limited number of barcodes (e.g., 96) available in commercial kits, we recommend 96 samples as a current practical maximum for multiplex plastome sequencing. This high-throughput approach should facilitate large-scale plastid genome sequencing at any level of phylogenetic diversity in angiosperms.</p>
</div>
</front>
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<affiliations><list><country><li>Royaume-Uni</li>
<li>République populaire de Chine</li>
<li>États-Unis</li>
</country>
<region><li>Angleterre</li>
<li>Angleterre de l'Est</li>
<li>Floride</li>
<li>Ohio</li>
<li>Zhejiang</li>
</region>
<settlement><li>Cambridge</li>
<li>Hangzhou</li>
</settlement>
<orgName><li>Université de Cambridge</li>
<li>Université de Zhejiang</li>
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</list>
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<name sortKey="Soltis, Douglas E" sort="Soltis, Douglas E" uniqKey="Soltis D" first="Douglas E." last="Soltis">Douglas E. Soltis</name>
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<name sortKey="Soltis, Pamela S" sort="Soltis, Pamela S" uniqKey="Soltis P" first="Pamela S." last="Soltis">Pamela S. Soltis</name>
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<name sortKey="Stull, Gregory W" sort="Stull, Gregory W" uniqKey="Stull G" first="Gregory W." last="Stull">Gregory W. Stull</name>
</country>
<country name="République populaire de Chine"><region name="Zhejiang"><name sortKey="Qi, Xinshuai" sort="Qi, Xinshuai" uniqKey="Qi X" first="Xinshuai" last="Qi">Xinshuai Qi</name>
</region>
</country>
<country name="Royaume-Uni"><region name="Angleterre"><name sortKey="Brockington, Samuel F" sort="Brockington, Samuel F" uniqKey="Brockington S" first="Samuel F." last="Brockington">Samuel F. Brockington</name>
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</tree>
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