Hybrid selection of discrete genomic intervals on custom-designed microarrays for massively parallel sequencing View Full Text


Ontology type: schema:ScholarlyArticle      Open Access: True


Article Info

DATE

2009-06

AUTHORS

Emily Hodges, Michelle Rooks, Zhenyu Xuan, Arindam Bhattacharjee, D Benjamin Gordon, Leonardo Brizuela, W Richard McCombie, Gregory J Hannon

ABSTRACT

Complementary techniques that deepen information content and minimize reagent costs are required to realize the full potential of massively parallel sequencing. Here, we describe a resequencing approach that directs focus to genomic regions of high interest by combining hybridization-based purification of multi-megabase regions with sequencing on the Illumina Genome Analyzer (GA). The capture matrix is created by a microarray on which probes can be programmed as desired to target any non-repeat portion of the genome, while the method requires only a basic familiarity with microarray hybridization. We present a detailed protocol suitable for 1-2 microg of input genomic DNA and highlight key design tips in which high specificity (>65% of reads stem from enriched exons) and high sensitivity (98% targeted base pair coverage) can be achieved. We have successfully applied this to the enrichment of coding regions, in both human and mouse, ranging from 0.5 to 4 Mb in length. From genomic DNA library production to base-called sequences, this procedure takes approximately 9-10 d inclusive of array captures and one Illumina flow cell run. More... »

PAGES

960-974

Identifiers

URI

http://scigraph.springernature.com/pub.10.1038/nprot.2009.68

DOI

http://dx.doi.org/10.1038/nprot.2009.68

DIMENSIONS

https://app.dimensions.ai/details/publication/pub.1009828502

PUBMED

https://www.ncbi.nlm.nih.gov/pubmed/19478811


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