55. Optical Mapping: A Complete System For Whole Genome Shotgun Mapping 

D.C. Schwartz^1,2, T. Anantharaman², J. Apodaca¹, C. Aston¹, V. Clarke¹, D. Gebauer¹, S. Delobette¹, E. Dimalanta¹, J. Edington¹, A. Evenzehav¹, J. Giacalone¹, V. Gibaja¹, C. Hiort¹, E. Huff¹, J. Jing¹, Z. Lai¹, D. Lazaro¹, E. Lee¹, J. Lin¹, K. Lin¹, B. Mishra², L. Ni¹, S. Paxia², B. Porter¹, R. Qi¹, A. Ramanathan¹, Y. Skiadis¹, J. Vafai¹, W. Wang¹, H. Zhao^1 
1W. M. Keck Laboratory for Biomolecular Imaging, Department of Chemistry, and ²Courant Institute of Mathematical Sciences, Department of Computer Science, New York University, NY 10003 
schwad01@mcrcr6.med.nyu.edu 

Optical Mapping is a single molecule approach for the rapid production of ordered restriction maps from individual DNA molecules. Fluorescence microscopy is used to directly image individual DNA molecules bound to derivatized glass surfaces, and cleaved by restriction enzymes. Fragments retain their original order, and cut sites are flagged by small, visible gaps. The Optical Mapping system has advanced in several critical areas to emerge as a means for the detailed mapping of both clones and entire genomes (Deinococcus radiodurans and Plasmodium falciparum). We mapped these entire microbial genomes using megabased-sized genomic DNA molecules (600-10,000 kb). Because large fragments of randomly sheared DNA are mapped with high cutting efficiency, many overlapping restriction site landmarks allow contigs to be assembled and a shotgun mapping strategy can be employed. High resolution whole genome maps can therefore be assembled without library construction and associated cloning artifacts. Because ensembles of single molecules are analyzed, small amounts of starting material are required enabling mapping of microorganisms, which are problematic to culture. Whole genome maps enable the size of the genome to be accurately determined, an important prelude to any sequencing endeavor. Most importantly, whole genome maps from genomic DNA provide an in situ picture of the architecture of the entire genome, revealing the number of chromosomes, existence of extrachromosomal elements etc. Populations can be potentially be characterized by comparing maps from different strains. Recent efforts have been to create high resolution maps of E. coli O157 strain (5.4 mgb) as a scaffold for facilitated sequence assembly and verification (Collaborator: F. Blattner, U. Wisconsin). We will compare maps generated "in silico" from the sequence of E. coli K12 (4.6 mgb) to identify regions that are unique to O157 and could be targeted for sequencing. Given the success we enjoyed in the restriction mapping of whole microbial genomes, and the proven reliability of the contig assembly algorithms developed for these efforts, we decided to construct a reference restriction map of the entire human genome. In four weeks our laboratory mapped 0.6 human genome equivalents at 40 kb resolution, using genomic fragments with average size of 2.1 mb. Our analysis of the contigs formed showed good correspondence with suitably modified Lander-Waterman physical mapping criteria in terms of the number and depth of overlapped genomic fragments. Goals are to simultaneously complete the human reference map to include 10-15x coverage and to link with other physical maps by the alignment of restriction mapped BAC contigs. The utility of this map will be to facilitate large scale sequencing projects and to provide a novel resource for the analysis of large populations.