Structural Variation Detection and De Novo Assembly in Complex Genomes Using Extremely Long Single-Molecule Imaging

De novo genome assemblies using only short read data are generally incomplete and highly fragmented due to the intractable complexity found in most genomes. This complexity, consisting mainly of large duplications and repetitive regions, hinders sequence assembly and subsequent comparative analyses.

We present a single molecule genome analysis system (Irys) based on NanoChannel Array technology that linearizes extremely long DNA molecules for observation. This high-throughput platform automates the imaging of single molecules of genomic DNA hundreds of kilobases in size to measure sufficient sequence uniqueness for unambiguous assembly of complex genomes. High-resolution genome maps assembled de novo from the extremely long single molecules retain the original context and architecture of the genome.  As a result, genome maps improve contiguity and accuracy of whole genome assemblies, permitting a more comprehensive analysis of functional genome biology and structural variation.

Additionally, genome maps serve as a much-needed orthogonal validation method to NGS assemblies. Free from reference bias, Genome Maps identify novel sequence insertions and locate transgene or viral integration sites.

In addition to providing an introduction to this newly available technology, we will demonstrate a number of examples of its utility in a variety of organisms, including examples from crop plants (wheat and rice), model species (Drosophila and Arabidopsis) and pest insects (spidermite). Genome maps are used to resolve repetitive functional elements and genes, validate or repair de novo sequence assemblies, and dissect cultivar differences and structural variants.