DOE Human Genome Program Contractor-Grantee Workshop VIII
February 27-March 2, 2000  Santa Fe, NM

52. Mapping and Functional Analysis of the Mouse Genome

D. K. Johnson,¹ C. T. Culiat,¹ M. L. Klebig,² Y. You,¹ D. R. Miller,¹ L. B. Russell,¹ E. J. Michaud,¹ and E. M. Rinchik^1,2

¹Mammalian Genetics and Development Section, Life Sciences Division, Oak Ridge National Laboratory, P.O. Box 2009, Oak Ridge, TN 37831-8077 and ²Department of Biochemistry, Molecular, and Cellular Biology, University of Tennessee, Knoxville, TN 37996

johnsondk@ornl.gov

As part of a functional-genomics strategy to determine how altered genes and proteins impact complex biological systems in mammals, the Mammalian Genetics Program at ORNL is characterizing a number of regions of the mouse genome on both the physical and functional levels, using mouse mutations as tools. This project forms a logical partnership with our regional mutagenesis program, which is designed to detect, maintain, and partially characterize new chemically induced mutations in ~8-10% of the mouse genome by utilizing new genetic tools and broad-based phenotype screening. The integrated efforts of these projects will advance the post-genome sequencing mission of annotating human DNA sequence with whole-organism functional information from the mouse model system.

Our goal is to acquire the DNA sequence of each region, to develop a validated transcription/ expression map, and to ascribe whole-organism functional information to each coding sequence through analysis of heritable gene mutations. Our chosen genome regions and subregions will be physically delimited by identifiable DNA landmarks (typically chromosomal rearrangements); hence, we can easily co-map mutant phenotypes with coding units to establish unambiguous sequence/function relationships by superimposing mutation maps onto transcription maps. The target regions include the 5- to 6-cM pink-eyed dilution (p) region in mouse Chromosome (Chr) 7 (human Chrs 11p, 15p, and 15q homologies); the 14 cM between p and the albino (Tyr; c) region (human Chr 15q); the 6- to 11-cM Tyr region (human Chrs 6p, 11p, 11q, and 15q); all of Chr 15 (human Chrs 5p, 8q, 12q, 22q), concentrating initially on the distal half; and mid-Chr 10 (human Chrs 6q, 10q, 12q, 21q, and 22q). With available molecular and embryonic stem (ES)-cell techniques, the growing emphasis on regional-mutagenesis strategies and the development of mouse reagents with which to carry out those strategies, we and others can extend this same discovery approach to any genome region.

Complete DNA sequence for these regions will be obtained by collaboration with the Joint Genome Institute or by mining of public databases created by the NIH mouse sequencing efforts. After ascer-tainment of potential transcription units from EST mapping and from computational analysis of raw DNA sequence by ORNL's Computational Biosciences Section, predicted transcription units will be verified by RNA analyses (Northerns, RT-PCR, RNase protection, and/or microarray procedures). The ultimate correlation of dense mutation maps with the transcription/expression maps has begun by identifying candidate mutant genes bearing ENU mutations, using densely mutagenized regions within the p- and Tyr regions as initial models with which to develop efficient mutation-scanning techniques. Phenotype gaps can also be filled with knockout/ gene-trap mutations for genes discovered in DNA sequence analysis but not represented as ENU mutations. All new DNA sequence information, expression information, and mutations will be advertised to interested partners via the WWW.

[Research sponsored by the Office of Biological and Environmental Research, USDOE, under contract DE-AC05-960R22464 with Lockheed Martin Energy Research, Inc.]