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Mobile element scanning (ME-Scan) by targeted high-throughput sequencing

BACKGROUND: Mobile elements (MEs) are diverse, common and dynamic inhabitants of nearly all genomes. ME transposition generates a steady stream of polymorphic genetic markers, deleterious and adaptive mutations, and substrates for further genomic rearrangements. Research on the impacts, population d...

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Autores principales: Witherspoon, David J, Xing, Jinchuan, Zhang, Yuhua, Watkins, W Scott, Batzer, Mark A, Jorde, Lynn B
Formato: Texto
Lenguaje:English
Publicado: BioMed Central 2010
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2996938/
https://www.ncbi.nlm.nih.gov/pubmed/20591181
http://dx.doi.org/10.1186/1471-2164-11-410
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author Witherspoon, David J
Xing, Jinchuan
Zhang, Yuhua
Watkins, W Scott
Batzer, Mark A
Jorde, Lynn B
author_facet Witherspoon, David J
Xing, Jinchuan
Zhang, Yuhua
Watkins, W Scott
Batzer, Mark A
Jorde, Lynn B
author_sort Witherspoon, David J
collection PubMed
description BACKGROUND: Mobile elements (MEs) are diverse, common and dynamic inhabitants of nearly all genomes. ME transposition generates a steady stream of polymorphic genetic markers, deleterious and adaptive mutations, and substrates for further genomic rearrangements. Research on the impacts, population dynamics, and evolution of MEs is constrained by the difficulty of ascertaining rare polymorphic ME insertions that occur against a large background of pre-existing fixed elements and then genotyping them in many individuals. RESULTS: Here we present a novel method for identifying nearly all insertions of a ME subfamily in the whole genomes of multiple individuals and simultaneously genotyping (for presence or absence) those insertions that are variable in the population. We use ME-specific primers to construct DNA libraries that contain the junctions of all ME insertions of the subfamily, with their flanking genomic sequences, from many individuals. Individual-specific "index" sequences are designed into the oligonucleotide adapters used to construct the individual libraries. These libraries are then pooled and sequenced using a ME-specific sequencing primer. Mobile element insertion loci of the target subfamily are uniquely identified by their junction sequence, and all insertion junctions are linked to their individual libraries by the corresponding index sequence. To test this method's feasibility, we apply it to the human AluYb8 and AluYb9 subfamilies. In four individuals, we identified a total of 2,758 AluYb8 and AluYb9 insertions, including nearly all those that are present in the reference genome, as well as 487 that are not. Index counts show the sequenced products from each sample reflect the intended proportions to within 1%. At a sequencing depth of 355,000 paired reads per sample, the sensitivity and specificity of ME-Scan are both approximately 95%. CONCLUSIONS: Mobile Element Scanning (ME-Scan) is an efficient method for quickly genotyping mobile element insertions with very high sensitivity and specificity. In light of recent improvements to high-throughput sequencing technology, it should be possible to employ ME-Scan to genotype insertions of almost any mobile element family in many individuals from any species.
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spelling pubmed-29969382010-12-07 Mobile element scanning (ME-Scan) by targeted high-throughput sequencing Witherspoon, David J Xing, Jinchuan Zhang, Yuhua Watkins, W Scott Batzer, Mark A Jorde, Lynn B BMC Genomics Methodology Article BACKGROUND: Mobile elements (MEs) are diverse, common and dynamic inhabitants of nearly all genomes. ME transposition generates a steady stream of polymorphic genetic markers, deleterious and adaptive mutations, and substrates for further genomic rearrangements. Research on the impacts, population dynamics, and evolution of MEs is constrained by the difficulty of ascertaining rare polymorphic ME insertions that occur against a large background of pre-existing fixed elements and then genotyping them in many individuals. RESULTS: Here we present a novel method for identifying nearly all insertions of a ME subfamily in the whole genomes of multiple individuals and simultaneously genotyping (for presence or absence) those insertions that are variable in the population. We use ME-specific primers to construct DNA libraries that contain the junctions of all ME insertions of the subfamily, with their flanking genomic sequences, from many individuals. Individual-specific "index" sequences are designed into the oligonucleotide adapters used to construct the individual libraries. These libraries are then pooled and sequenced using a ME-specific sequencing primer. Mobile element insertion loci of the target subfamily are uniquely identified by their junction sequence, and all insertion junctions are linked to their individual libraries by the corresponding index sequence. To test this method's feasibility, we apply it to the human AluYb8 and AluYb9 subfamilies. In four individuals, we identified a total of 2,758 AluYb8 and AluYb9 insertions, including nearly all those that are present in the reference genome, as well as 487 that are not. Index counts show the sequenced products from each sample reflect the intended proportions to within 1%. At a sequencing depth of 355,000 paired reads per sample, the sensitivity and specificity of ME-Scan are both approximately 95%. CONCLUSIONS: Mobile Element Scanning (ME-Scan) is an efficient method for quickly genotyping mobile element insertions with very high sensitivity and specificity. In light of recent improvements to high-throughput sequencing technology, it should be possible to employ ME-Scan to genotype insertions of almost any mobile element family in many individuals from any species. BioMed Central 2010-06-30 /pmc/articles/PMC2996938/ /pubmed/20591181 http://dx.doi.org/10.1186/1471-2164-11-410 Text en Copyright ©2010 Witherspoon et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Methodology Article
Witherspoon, David J
Xing, Jinchuan
Zhang, Yuhua
Watkins, W Scott
Batzer, Mark A
Jorde, Lynn B
Mobile element scanning (ME-Scan) by targeted high-throughput sequencing
title Mobile element scanning (ME-Scan) by targeted high-throughput sequencing
title_full Mobile element scanning (ME-Scan) by targeted high-throughput sequencing
title_fullStr Mobile element scanning (ME-Scan) by targeted high-throughput sequencing
title_full_unstemmed Mobile element scanning (ME-Scan) by targeted high-throughput sequencing
title_short Mobile element scanning (ME-Scan) by targeted high-throughput sequencing
title_sort mobile element scanning (me-scan) by targeted high-throughput sequencing
topic Methodology Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2996938/
https://www.ncbi.nlm.nih.gov/pubmed/20591181
http://dx.doi.org/10.1186/1471-2164-11-410
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