U.S. Department of Energy

Human Genome 1993 Program Report: Research Centers

Date Published: March 1994

Established by DOE to foster collaborations by teams of investigators from various disciplines, the three DOE human genome research centers [located at Lawrence Berkeley Laboratory (LBL), Lawrence Livermore National Laboratory (LLNL), and Los Alamos National Laboratory (LANL)] address such major tasks as genetic and physical mapping, DNA sequencing, informatics related to mapping and sequencing, and technology development. Contracts to these centers are funded annually and peer reviewed through site visits every 2 to 3 years. In FY 1993, $30.365 million (48.8%) of the DOE OHER genome program budget was devoted to the centers, with LLNL receiving $9.713 million, LBL $8.599 million, and LANL $12.053 million.

Centers also play a key role in promoting distribution of genome research technology and resources through outside collaborations, public access to laboratory databases, and "visitor laboratories" at which visiting scientists and pre- and postdoctoral students can apply genome center expertise and technology to their own research. In addition, the centers strive to make available their resources--including biologicals, software, databases, instrumentation, and training opportunities--to the entire genome research community.

All center investigators are encouraged to engage in active collaborations with the private sector and transfer their resources and technologies for commercial development. Activities include the transfer of vectors, primers, and software to industry and the further development of instrumentation with industrial partners. Short descriptions and recent accomplishments of each center follow.

Lawrence Berkeley Laboratory

Established in 1988, the LBL Human Genome Center is developing research and analytical technology to speed genome mapping and sequencing and decrease costs. Research at the LBL center focuses on mapping and sequencing chromosome 21, which is the smallest human chromosome and contains an estimated 900 genes. Identified chromosome 21 genes include those implicated in Down's syndrome and familial Alzheimer's disease. A major goal of the center is to use low-density maps to catalyze construction of high-density genetic and physical maps, gene maps, and sequence. Toward this goal the center is sequencing a biologically interesting 3- to 4-Mb region of chromosome 21 (including the Down's syndrome region) and creating high-density genetic maps of that chromosome. Center investigators are aided by groups specializing in robotics, instrumentation automation, and informatics development. The current focus at LBL is to develop technology to sequence 10 Mb or more per year at $0.50 per base with more than 99.9% accuracy. In 1993 Jasper Rine was the center's director, and Sylvia Spengler was the deputy director. In January 1994 Mohandas Narla became acting director of the Human Genome Center.

Recent Accomplishments

• An 80-kb insert from a P1 clone from the Down's region was sequenced using the DOG-tag strategy that generates distance, orientation, and gene-size resolution. Over 150 P1 clones covering the Down's region have been selected and mapped to yeast artificial chromosomes (YACs). These P1s will provide substrates for the DOG-tag strategy.
  
• Novel mapping resources were developed for chromosome 21, including 60 mapped cDNA clones, over 40 dinucleotide repeat-based genetic markers, a YAC contig map, a set of somatic cell radiation hybrids (with David Cox, Stanford University), a cytogenetic map of the long arm enriched by fluorescence in situ hybridization (FISH) with 280 YACs and cosmids, and 188 cosmids forming 21 multiple YAC contigs and 4 single YAC contigs.
  
• Instruments supporting automated mapping and sequencing were developed, including an automated colony picker and imaging station with supporting hardware for gel casting and an expandable high-throughput polymerase chain reaction (PCR) apparatus.
  
• Companion software produced includes 21Bdb, a database variant of ACEDB (the Caenorhabditis elegans database) containing physical mapping and sequence data for chromosome 21; a sequence-analysis package; database management tools, including ERDRAW, SDT, QST, and OPM Editor; and an automated generator of user interfaces that changes the interface in response to changes in the underlying database.

Lawrence Livermore National Laboratory

The Human Genome Center at LLNL was established in 1990 as an outgrowth of studies on the identification and characterization of human DNA repair genes, specifically on chromosome 19. Major goals include new cloning, mapping, instrumentation, informatics, and sequencing technologies focused on the assembly, closure, and characterization of a high-resolution ordered clone map of human chromosome 19. The final high-resolution map will consist of cosmid contigs with YACs, bacterial artificial chromosomes (BACs), and P1 artificial chromosomes (PACs), as well as an EcoR1 restriction map for the minimal spanning set of cosmids. The physical map is being aligned with genetic maps of chromosome 19. Other goals are to isolate, map, and sequence chromosome 19 cDNAs, with emphasis on full-length clones; construct (with LANL) National Laboratory Gene Library Project (NLGLP) chromosome-specific lambda and cosmid libraries for distribution; and develop new cloning, mapping, and sequencing technologies. Anthony V. Carrano is the center's director.

Recent Accomplishments

• Coverage has been achieved on an estimated 90% of chromosome 19 with about 800 contigs assembled from over 12,000 cosmids, with use of automated fluorescence-based fingerprinting.
  
• Rapid closure of gaps is progressing with the use of YACs, BACs, and PACs; 400 contigs representing about 60% of the chromosome have been regionally mapped to bands by FISH.
  
• More than 180 genes and polymorphic markers were localized on the contig map. FISH was used to localize over 800 cosmids to bands, including an ordered set of cosmids spaced an average of 1 Mb over the whole chromosome and 500 kb in selected bands.
  
• Over 166 kb from chromosome 19 repair-gene regions were sequenced.
  
• With several collaborators, organization was determined for a number of gene families, including carcinoembryonic, olfactory receptor, zinc finger, cytochrome P450, and fucosyl transferase.
  
• New vectors for cosmid and P1 cloning systems and novel Alu PCR primers for fingerprinting and forensic analysis were developed.
  
• Construction was completed for the NLGLP chromosome-specific lambda and cosmid libraries from sorted chromosomes 3, 7, and X; libraries for chromosomes 9, 12, 18, 19, 21, 22, and Y are also complete.
  
• Integrated mapping-analysis software was developed with interactive graphical display and linkage to local and national databases.
  
• A high-speed flow cytometer with sorting capabilities was completed for cell and chromosome purification.
  
• New instrumentation was developed for high-density filter production.

Contacts: Linda Ashworth, Assistant to Center Director (510/422-5665, ashworth1@ llnl.gov) or Anthony Carrano, Director (510/422-5698, Fax: /423-3110, carrano1@llnl.gov); Human Genome Center; LLNL; Biology and Biotechnology Research Program; 7000 East Avenue, L-452; P.O. Box 808; Livermore, CA 94551.

Graduate and postdoctoral research training is available through the Institute of Genetics and Genomics at LLNL (Harvey Mohrenweiser, 510/423-0534).

Los Alamos National Laboratory

The Center for Human Genome Studies at LANL was established in 1988. The LANL center's goals include assembly of complete high-resolution (0.1 Mb) maps of chromosome 16 and regions of chromosome 5, studies at the molecular level of chromosome structure and function, and isolation of selected genes of interest on chromosomes 5 and 16. Other goals are (1) short-term computational development and support for large-scale physical mapping and sequencing projects and long-term development of tools for storage, manipulation, and analysis of genome data; (2) development and application of new methods for physical mapping; (3) use of robotics in handling and storing DNA fragments; (4) construction of DNA libraries from flow-sorted chromosomes; (5) rapid, inexpensive, large-scale sequencing; and (6) studies of ethical, legal, and social issues arising from the increased availability and use of genome data. Technology transfer activities are progressing and include robotic instrumentation development, software licensing, library distribution, and rapid sequencing technology. Robert K. Moyzis is the center's director and Larry L. Deaven is the deputy director.

Recent Accomplishments

• Two maps have been constructed: (1) a high-resolution cosmid/YAC physical map of human chromosome 16 consisting of regionally localized contigs covering over 95% of the chromosome and (2) a framework sequence tagged site map of human chromosome 5 consisting of 300 new markers regionally assigned with a resolution of 4 Mb on 5q to 1 Mb on 5p.
  
• The unusual 3-D structure of telomeric DNA was determined (with Alex Rich, Massachusetts Institute of Technology); the human telomere represents the endpoint for genetic and physical maps and was identified and cloned at LANL in 1988.
  
• Highly conserved centromeric repetitive DNA regions, likely human centromere components, were identified and cloned.
  
• A novel complementary DNA (cDNA) library was constructed from mRNA obtained from a paternally encoded human pregnancy (hydatidiform mole).
  
• NLGLP chromosome-specific phage and cosmid libraries were constructed with LLNL (over 2500 DNA libraries were sent to research and production laboratories worldwide, including complete digest libraries for each human chromosome); also constructed were partial-digest phage or cosmid libraries for human chromosomes 4, 5, 6, 8, 10, 11, 13, 14, 16, 17, 20, X, and Y; and complete digest low-chimeric YAC libraries for human chromosomes 5, 9, 16, and 21.
  
• Flow-cytometry techniques were developed to detect single DNA molecules, resulting in a Cooperative Research and Development Agreement (called CRADA) with Life Technologies, Inc., for codevelopment of a rapid DNA-sequencing technology.
  
• A robot was constructed for high-density cosmid/YAC array replication and distribution.
  
• SIGMA (System for Integrated Genome Map Assembly), an X windows-based software tool, was developed for creating, editing, and viewing integrated genome maps.
  
• cDNA-Inform, a collection of public and private sequences and software for automatic searches and comparisons, was produced.

Contact: Lynn Clark, Technical Coordinator (505/667-9376, Fax: -2891; moyzis@ flovax.lanl.gov); Center for Human Genome Studies; LANL; Life Sciences Division, MS M886; Los Alamos, NM 87545.