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Solution-based targeted genomic enrichment for precious DNA samples

BACKGROUND: Solution-based targeted genomic enrichment (TGE) protocols permit selective sequencing of genomic regions of interest on a massively parallel scale. These protocols could be improved by: 1) modifying or eliminating time consuming steps; 2) increasing yield to reduce input DNA and excessi...

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Detalles Bibliográficos
Autores principales: Shearer, Aiden Eliot, Hildebrand, Michael S, Smith, Richard JH
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3561655/
https://www.ncbi.nlm.nih.gov/pubmed/22559009
http://dx.doi.org/10.1186/1472-6750-12-20
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author Shearer, Aiden Eliot
Hildebrand, Michael S
Smith, Richard JH
author_facet Shearer, Aiden Eliot
Hildebrand, Michael S
Smith, Richard JH
author_sort Shearer, Aiden Eliot
collection PubMed
description BACKGROUND: Solution-based targeted genomic enrichment (TGE) protocols permit selective sequencing of genomic regions of interest on a massively parallel scale. These protocols could be improved by: 1) modifying or eliminating time consuming steps; 2) increasing yield to reduce input DNA and excessive PCR cycling; and 3) enhancing reproducible. RESULTS: We developed a solution-based TGE method for downstream Illumina sequencing in a non-automated workflow, adding standard Illumina barcode indexes during the post-hybridization amplification to allow for sample pooling prior to sequencing. The method utilizes Agilent SureSelect baits, primers and hybridization reagents for the capture, off-the-shelf reagents for the library preparation steps, and adaptor oligonucleotides for Illumina paired-end sequencing purchased directly from an oligonucleotide manufacturing company. CONCLUSIONS: This solution-based TGE method for Illumina sequencing is optimized for small- or medium-sized laboratories and addresses the weaknesses of standard protocols by reducing the amount of input DNA required, increasing capture yield, optimizing efficiency, and improving reproducibility.
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spelling pubmed-35616552013-02-05 Solution-based targeted genomic enrichment for precious DNA samples Shearer, Aiden Eliot Hildebrand, Michael S Smith, Richard JH BMC Biotechnol Methodology Article BACKGROUND: Solution-based targeted genomic enrichment (TGE) protocols permit selective sequencing of genomic regions of interest on a massively parallel scale. These protocols could be improved by: 1) modifying or eliminating time consuming steps; 2) increasing yield to reduce input DNA and excessive PCR cycling; and 3) enhancing reproducible. RESULTS: We developed a solution-based TGE method for downstream Illumina sequencing in a non-automated workflow, adding standard Illumina barcode indexes during the post-hybridization amplification to allow for sample pooling prior to sequencing. The method utilizes Agilent SureSelect baits, primers and hybridization reagents for the capture, off-the-shelf reagents for the library preparation steps, and adaptor oligonucleotides for Illumina paired-end sequencing purchased directly from an oligonucleotide manufacturing company. CONCLUSIONS: This solution-based TGE method for Illumina sequencing is optimized for small- or medium-sized laboratories and addresses the weaknesses of standard protocols by reducing the amount of input DNA required, increasing capture yield, optimizing efficiency, and improving reproducibility. BioMed Central 2012-05-04 /pmc/articles/PMC3561655/ /pubmed/22559009 http://dx.doi.org/10.1186/1472-6750-12-20 Text en Copyright ©2012 Shearer et al.; licensee BioMed Central Ltd. 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
Shearer, Aiden Eliot
Hildebrand, Michael S
Smith, Richard JH
Solution-based targeted genomic enrichment for precious DNA samples
title Solution-based targeted genomic enrichment for precious DNA samples
title_full Solution-based targeted genomic enrichment for precious DNA samples
title_fullStr Solution-based targeted genomic enrichment for precious DNA samples
title_full_unstemmed Solution-based targeted genomic enrichment for precious DNA samples
title_short Solution-based targeted genomic enrichment for precious DNA samples
title_sort solution-based targeted genomic enrichment for precious dna samples
topic Methodology Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3561655/
https://www.ncbi.nlm.nih.gov/pubmed/22559009
http://dx.doi.org/10.1186/1472-6750-12-20
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