e-journal

Impact of enrichment conditions on cross-species capture of fresh and degraded DNA

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By combining high-throughput sequencing with target enrichment (‘hybridization capture’), researchers are able to
obtain molecular data from genomic regions of interest for projects that are otherwise constrained by sample quality
(e.g. degraded and contamination-rich samples) or a lack of a priori sequence information (e.g. studies on nonmodel
species). Despite the use of hybridization capture in various fields of research for many years, the impact of enrichment conditions on capture success is not yet thoroughly understood. We evaluated the impact of a key parameter –
hybridization temperature – on the capture success of mitochondrial genomes across the carnivoran family Felidae.
Capture was carried out for a range of sample types (fresh, archival, ancient) with varying levels of sequence divergence between bait and target (i.e. across a range of species) using pools of individually indexed libraries on Agilent SureSelectTM arrays. Our results suggest that hybridization capture protocols require specific optimization for the sample type that is being investigated. Hybridization temperature affected the proportion of on-target sequences following capture: for degraded samples, we obtained the best results with a hybridization temperature of 65 °C, while
a touchdown approach (65 °C down to 50 °C) yielded the best results for fresh samples. Evaluation of capture performance at a regional scale (sliding window approach) revealed no significant improvement in the recovery of DNA
fragments with high sequence divergence from the bait at any of the tested hybridization temperatures, suggesting
that hybridization temperature may not be the critical parameter for the enrichment of divergent fragments.

Keywords: degraded DNA, Felidae, hybridization capture, mitogenomes, next-generation sequencing, sequence enrichment
Received 3 December 2014; revision received 21 April 2015; accepted 23 April 2015

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Informasi Detail

Judul Seri

Molecular Ecology Resources

No. Panggil

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Penerbit

: John Wiley & Sons Ltd., 2016

Deskripsi Fisik

Molecular Ecology Resources (2016) 16, 42–55

Bahasa

English

ISBN/ISSN

doi: 10.1111/1755-09

Klasifikasi

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Tipe Isi

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Tipe Media

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Tipe Pembawa

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Edisi

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Subjek

BIOLOGI

Info Detail Spesifik

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Pernyataan Tanggungjawab

Wati/Agus

Versi lain/terkait

Lampiran Berkas

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