TY - JOUR T1 - SHIMS 3.0: Highly efficient single-haplotype iterative mapping and sequencing using ultra-long nanopore reads JF - bioRxiv DO - 10.1101/2020.09.18.303735 SP - 2020.09.18.303735 AU - Daniel W. Bellott AU - Ting-Jan Cho AU - Emily K. Jackson AU - Helen Skaletsky AU - Jennifer F. Hughes AU - David C. Page Y1 - 2020/01/01 UR - http://biorxiv.org/content/early/2020/09/20/2020.09.18.303735.abstract N2 - The reference sequence of structurally complex regions can only be obtained through a highly accurate clone-based approach that we call Single-Haplotype Iterative Mapping and Sequencing (SHIMS). In recent years, improvements to SHIMS have reduced the cost and time required by two orders of magnitude, but internally repetitive clones still require extensive manual effort to transform draft assemblies into reference-quality finished sequences. Here we introduce SHIMS 3.0, using ultra-long nanopore reads to resolve internally repetitive structures and minimize the need for manual finishing of Illumina-based draft assemblies. This protocol proceeds from clone-picking to finished assemblies in 2 weeks for about 80 dollars per clone. We have used SHIMS 3.0 to finish the structurally complex TSPY array on the human Y chromosome, which could not be resolved by previous sequencing methods. Our protocol provides access to structurally complex regions that would otherwise be inaccessible from whole-genome shotgun data or require an impractical amount of manual effort to generate an accurate assembly.Competing Interest StatementThe authors have declared no competing interest. ER -