ANNUAL PROGRESS REPORT

NATIONAL RESEARCH SUPPORT PROJECT — NRSP-8

Year Ending 2002

Preliminary Information — Not for Publication

Michigan Station Report to the NRSP-8 Swine Genome Committee

Submitted by Catherine W. Ernst

  1. PROJECT TITLE:
    2.

    National Animal Genome Research Program (NAGRP)

  2. COOPERATING AGENCIES AND PRINCIPAL LEADERS:
  1. Agencies and Departments: Michigan Agricultural Experiment Station and Department of Animal Science, Michigan State University
  2. Project Leader: Catherine Ernst
  3. Cooperating Investigators: N.E. Raney, V.D. Rilington, P.J. Venta, D. Housley, R.O. Bates, M.E. Doumit, D. Edwards and M. Hoge, Michigan State University; G.A. Rohrer, USDA-ARS, US Meat Animal Research Center; M.F. Rothschild, U.S. Swine Genome Coordinator
  1. NATURE OF WORK AND PRINCIPAL RESULTS OF YEAR:

Objective 1. Develop high resolution comparative genome maps aligned across species that link agricultural animal maps to those of the human and mouse genomes.

The objective of an ongoing project in our laboratory is to further develop the pig comparative map by identifying pig sequence-tagged sites (STS) for human genes and mapping each STS in the pig by physical assignment, RH mapping and/or genetic linkage analysis. Comparative mapping allows transfer of information from marker-rich genomes such as the human to species with limited mapping resources. Current pig genome maps contain relatively few Type I (gene) markers and mapping of such markers will clarify gene order and improve the utility of comparative maps. The need for identifying additional polymorphism in Type I markers for genetic linkage mapping can be fulfilled by detection of single-nucleotide polymorphisms (SNPs). For SNP identification, we use a pool-and-sequence strategy in which a pool of DNA samples from different breeds is PCR-amplified and sequenced. SNPs are identified as ambiguous bands occurring at identical positions in a sequencing gel. We have developed 133 pig STS, placed 62 STS on a pig radiation hybrid (RH) map using the INRA-University of Minnesota porcine RH panel and identified 110 SNPs in 40 STS (1-6 SNPs per STS). PCR primers were designed using heterologous sequences and the identity of each STS was confirmed by DNA sequencing. Two-point analysis of RH data showed significant linkage of STS with markers on 12 different pig chromosomes. Genetic linkage analysis has been performed using SNP genotypes for 7 STS in the PiGMaP reference families and 7 additional STS in the MARC reference families. Results from this study will help to further improve the pig and human comparative map.

Objective 2. Increase the marker density of existing linkage maps used in QTL mapping and integrate them with physical maps of animal chromosomes.

Development of a Duroc X Pietrain pig resource population was begun in 2000. The foundation of the population consisted of HAL 1843TM free Pietrain females and Duroc males. Twenty HAL 1843TM free Pietrain females were purchased in January, 2000. Of these, 16 farrowed litters with 124 pigs weaned. Three boars per foundation Duroc sire and 60 females were retained from F1 litters for breeding purposes. All other F1 animals were slaughtered and carcass measurements and meat quality characteristics determined. From the summary of meat quality characteristics, sons from the two most desirable foundation Duroc sires were retained to produce the F2 generation. F1 boars were trained for semen collection for artificial insemination. The F1 breeding animals are currently being inter se mated with the goal of producing 1,000 F2 progeny. Data being collected includes the following growth characteristics: birth weight, weaning weight, 10 week weight and weight before slaughter near 113 kg. At slaughter, further data collection includes: carcass weight and pH45min. After 24 hr chilling, subsequent data collection includes: carcass backfat depth (1st rib, last rib, last lumbar, tenth rib, off midline), loin muscle area and carcass length. Carcasses are processed and closely trimmed wholesale cut weights of the ham, loin, belly, Boston butt and picnic are recorded. A loin section from the tenth to last rib is partitioned into 2.54 cm chops with the following data collected: NPPC color, firmness and marbling scores, Japanese color scores, Minolta color measurements, pHu, 24 hour post-processing drip loss, cooking loss, Warner Bratzler shear force values and intramuscular lipid. In addition, samples of loin muscle, backfat and liver are being collected at slaughter from a subset of F2 animals for use in gene expression analyses. Using DNA samples taken from the founder animal s along with the F1 and F2 pigs, segregating polymorphisms at microsatellite marker loci will be identified. Marker loci genotypes will be compared with phenotypic data to determine if there are significant phenotypic differences among animals with different allelic complements. White blood cells have been or will be collected from all F0, F1 and F2 individuals and are being stored for future DNA isolation. As of January 2003, 570 F2 offspring have reached market weight and been harvested. Production of F2 animals and collection of phenotypic data are expected to be completed by January of 2004.
  1. APPLICATION OF FINDINGS:

Results of this work will improve the pig comparative map. In addition, genetic linkage mapping of additional Type I markers will help to increase the marker density of existing linkage maps that are used in QTL mapping. Also, development of a unique pig resource population will provide a novel resource for identifying QTL associated with growth and carcass merit in pigs.

  1. WORK PLANNED FOR NEXT YEAR:

Comparative mapping of Type I loci will continue. Pig STS will be mapped using a radiation hybrid panel (Yerle et al., 1998; Hawken et al., 1999). Single-nucleotide polymorphisms (SNP) will be identified in pig STS and will be used for genetic linkage mapping. In addition, breeding and data collection will continue for the pig resource population.
  1. PUBLICATIONS FOR THE YEAR:

Yao, J., P.M. Coussens, P. Saama, S. Suchyta and C.W. Ernst. 2002. Generation of expressed sequence tags from a normalized porcine skeletal muscle cDNA library. Anim. Biotech. 13:211-202.

Ernst, C.W., V.D. Rilington, N.E. Raney, J. Yao, S.S. Sipkovsky, P.M. Saama, R.J. Tempelman and P.M. Coussens. 2002. Use of cDNA microarrays to detect differentially expressed genes in developing pig skeletal muscle. Plant and Animal Genome X meetings. San Diego, CA. http://www.intl-pag.org/pag/10/abstracts/PAGX_P702.html.

Rilington, V.D., P.J. Venta, D. Housley, N.E. Raney, S.R. Wesolowski, C.P. Wilkinson and CW. Ernst. 2002. Radiation hybrid mapping of and identification of single nucleotide polymorphisms in pig sequence tagged-sites. Plant and Animal Genome X meetings. San Diego, CA. http://www.intl-pag.org/pag/10/abstracts/PAGX_P601.html.