P0103 New Sequence Variants Identified by Region-Specific Extraction

Johannes Dapprich , Generation Biotech, LAWRENCEVILLE, NJ
Kristen Updike , Generation Biotech, Lawrenceville, NJ
Next-generation sequencing and targeted enrichment have become mainstream methods used in current genetic research.  However, the enormous throughput of the various platforms as well as other advantages over conventional genotyping and sequencing methods cannot always resolve more complex sequence regions.  Problems typically occur when the genome of interest is highly polymorphic, polyploid, or when a reference sequence is incomplete, contains gaps of unknown sequence, or does not exist yet.  One of the causes is the difficulty of short-read technologies to assemble accurate linkage information and haploid sequence over distances that exceed several kb.  This creates a challenge for the accurate resolution of homologous regions, repeat elements, structural variations and new alleles.  Sequence associated with structural variations can be lost when using short-fragment-based enrichment methods and may not be revealed by paired-end sequencing strategies. Region- and haplotype-specific extraction (RSE/HSE) is an automated, magnetic particle-based target enrichment method that captures large (10-40kb) segments of genomic DNA with high efficiency in a sequence- or SNP-specific way.  It provides the ability to determine extended molecular sequence and haplotypes from each target point.  A target point can be any specific target sequence or a known polymorphic position.  RSE/HSE has been used for the resolution of complex genomic regions and ambiguous allele combinations, the identification of previously undetected structural variants, QTL and transposon / T-DNA insertion point mapping, and to determine breakpoints and unknown sequence surrounding a specific associated marker, such as a SNP.