title: Targeted sequencing of large genomic regions with CATCH-Seq.
abstract: Current target enrichment systems for large-scale next-generation sequencing typically require synthetic oligonucleotides used as capture reagents to isolate sequences of interest. The majority of target enrichment reagents are focused on gene coding regions or promoters en masse. Here we introduce development of a customizable targeted capture system using biotinylated RNA probe baits transcribed from sheared bacterial artificial chromosome clone templates that enables capture of large, contiguous blocks of the genome for sequencing applications. This clone adapted template capture hybridization sequencing (CATCH-Seq) procedure can be used to capture both coding and non-coding regions of a gene, and resolve the boundaries of copy number variations within a genomic target site. Furthermore, libraries constructed with methylated adapters prior to solution hybridization also enable targeted bisulfite sequencing. We applied CATCH-Seq to diverse targets ranging in size from 125 kb to 3.5 Mb. Our approach provides a simple and cost effective alternative to other capture platforms because of template-based, enzymatic probe synthesis and the lack of oligonucleotide design costs. Given its similarity in procedure, CATCH-Seq can also be performed in parallel with commercial systems.
authors.affiliation: HudsonAlpha Institute for Biotechnology, Huntsville, Alabama, United States of America.
authors.forename: Kenneth
authors.initials: K
authors.lastname: Day
authors.affiliation: HudsonAlpha Institute for Biotechnology, Huntsville, Alabama, United States of America.
authors.forename: Jun
authors.initials: J
authors.lastname: Song
authors.affiliation: HudsonAlpha Institute for Biotechnology, Huntsville, Alabama, United States of America.
authors.forename: Devin
authors.initials: D
authors.lastname: Absher
mesh_terms: D002880:Chromosomes, Human, Pair 11 / Q000235:genetics; D002999:Clone Cells; D056915:DNA Copy Number Variations; D019175:DNA Methylation / Q000235:genetics; D015894:Genome, Human; D006801:Humans; D009693:Nucleic Acid Hybridization / Q000379:methods*; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D017422:Sequence Analysis, DNA / Q000379:methods*; D013698:Templates, Genetic