Solution-based targeted genomic enrichment for precious DNA samples

Aiden Eliot Shearer,Richard Jh Smith,Michael S Hildebrand

doi:10.1186/1472-6750-12-20

Abstract

BackgroundSolution-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.ResultsWe 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.ConclusionsThis 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.

Highlights

Solution-based targeted genomic enrichment (TGE) protocols permit selective sequencing of genomic regions of interest on a massively parallel scale
We developed a solution-based Targeted genomic enrichment (TGE) method for downstream Illumina sequencing that incorporates improvements proposed by Fisher et al, 2011 and Mamanova et al, 2010, in a non-automated workflow
In all cases we add standard Illumina barcode indexes during the post-hybridization amplification to allow for sample pooling prior to sequencing

Summary

Introduction

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 reproducibility. Targeted genomic enrichment (TGE) approaches were developed to address these issues by reducing cost and variant analysis complexity by screening specific disease-associated genomic. Solid-phase TGE was developed first, unlike solution-based TGE it requires expensive hybridization equipment and is less scalable. What may be the primary consideration for a small-to-medium sized laboratory wishing to perform TGE may be the expense associated with ancillary equipment. The combination of scalability and little-to-no infrastructure investment has made solution-based TGE the preferred method for TGE in many laboratories

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Conclusion