This project develops a platform-independent target enrichment technology for the preparation of large (50- 150 kb+) DNA fragments. These are magnetically selected from genomic regions of interest and used directly - without any intermediate amplification step - for next-generation sequencing (NGS). This will allow the direct detection of methylated base information on several of the newer NGS systems. Large DNA templates can currently be processed by several NGS platforms, and even more powerful systems are being developed. The ability to utilize large, contiguous DNA segments as sequencing templates greatly assists in detecting and characterizing structural variants, obtaining accurate sequence information across complex and GC-rich genomic loci, conducting de novo sequence assembly, phasing long-range haplotype information, and finding new or unexpected sequence elements, such as viral integration sites, inversions, duplications and other types of chromosomal rearrangements. Existing methods for genomic enrichment are not able to provide a comprehensive characterization of complex genomic loci in an economical and flexible manner because they cannot provide the large size fragments that are required to successfully span confounding sequence elements and resolve persistent challenges at important loci. We will develop an efficient long-read capture technology for small amounts of input DNA that resolves common sequencing and assembly problems that occur with most current amplification- or hybridization- based enrichment technologies. The benefits of this large-fragment enrichment as carried out with capture primers with a very small footprint (20-25 bases) will extend to all types of NGS platforms, including to those that rely on short read lengths. We believe that we can significantly improve the ability of researchers and clinicians to obtain complete and accurate characterizations of complex genomic variations and regions of interest that are important for disease diagnosis and prognosis by NGS. Anonymous, pre-existing samples for this project are provided by the NCI.