WASHINGTON 2002 ANNUAL REPORT TO NRSP-8

Submitted by Zhihua Jiang

January 11, 2003

I. Project: NRSP-8: Swine Genome Committee

II. Cooperating Agencies and Principal Investigators

A. Agencies and Departments Cooperating: Washington Agriculture Experiment Station and Animal Sciences Department, Washington State University

B. Leader of the Project: Zhihua Jiang
  1. Objectives

Objective 1: Develop high resolution comparative genome maps aligned across species

that link agricultural animal maps to those of the human and mouse genomes.

Objective 2: Increase the marker density of existing linkage maps used in QTL mapping

and integrate them with physical maps of animal chromosomes.

Objective 3: Expand and enhance internationally shared species genome databases and

provide other common resources that facilitate genome mapping.

IV. General Project Plan (2002)

A. Map-based chromosomal painting of the porcine genome on the human genome (Objective 1)

Genome mapping in pigs has advanced significantly in recent years. We use a CSAM (contiguous sets of autosomal markers) approach to generate a map-based chromosomal painting of the porcine genome on the human genome. A CSAM is an uninterrupted set of markers in one genome (primary genome) that is syntenic in the other genome (secondary genome). The CSAM approach requires relative marker order information in one genome and only the chromosome number of those markers in the other genome. As relative orders of genes in the human genome are known for CSAM mapping, the human genome is treated as primary, while the porcine genome is treated as secondary.

    1. Toward a comprehensive comparative map of porcine chromosome 3 using orthologous genes as landmarks (Objective 1)

Measuring conservation of contiguous sets of autosomal markers between human and porcine genomes has revealed that SSC3 is homologous to the regions 0 - 155 Mb on HSA2, 0 - 12 Mb and 45 - 130 Mb on HSA7, and 0 - 50 Mb on HSA16. Eighty-seven orthologous genes were selected as landmarks at approximately 3 Mb intervals in order to cover these homologous human segments. However, these markers could not generate a comprehensive RH map for SSC3, but with 7 linkage groups. In order to fill the gaps between linkage groups, we propose to add 30 markers more to the map. All of these markers will be typed on a pig/hamster RH panel and a comprehensive comparative map of SSC3 will be constructed using RHMAP (3.0) program.

C. A virtual bridge between microsatellite markers and functional genes (Objective 2)

Microsatellite markers have played an important role in porcine genome and QTL mapping. We use two approaches to link the microasatellite markers to functional gene regions: identification of their orthologous regions on the human genome and use of their close-linked markers of functional genes. The analysis has made a preliminary map-based chromosomal painting of the human genome on the porcine genome.

D. Development of a livestock orthologous gene (LOG) database (Objective 3)

In livestock species, two leading species in EST sequencing are cattle and swine. At the moment, there are about 240,000 bovine and 115,000 porcine EST entries in the GenBank which were sequenced with cDNA libraries derived from single tissue or pooled tissues (data accessed on January 1, 2003). A large scale of EST sequencing in chicken, horse, dog and cat is under way. We have been developing a gene-oriented approach to annotate gene/EST sequences in livestock species. The major procedures of our approach include: BLAST searches against GenBank databases using human cDNA sequences as queries, identification of orthologous gene/EST segments in livestock species, generation of tentative or orthologous consensus sequences by assembling and ortholog assignment and comparative mapping.

V. Work Progress

    1. Map-based chromosomal painting of the porcine genome on the human genome (Objective 1)

Recently we updated the human-porcine genome painting based on a total of 1438 porcine genes/markers that were detected to have their orthologs in the human genome, including 730 known genes, 342 expressed sequence tags (ESTs), 173 sequence tagged sites (STSs) and 193 microsatellite markers. The painting revealed 124 homologous segments between human and porcine genomes, including 36 singletons.

B. Toward a comprehensive comparative map of porcine chromosome 3 using orthologous genes as landmarks (Objective 1)

Nineteen orthologous gene markers have been added to the map and the number of linkage groups is reduced from 7 to 4. The largest linkage group has 54 markers now.

C. A virtual bridge between microsatellite markers and functional genes (Objective 2)

A total of 469 microsatellite markers have been found to be linked to functional gene regions, including 42 on SSC1, 31 on SSC2, 29 on SSC3, 31 on SSC4, 24 on SSC5, 35 on SSC6, 21 on SSC7, 27 on SSC8, 24 on SSC9, 16 on SSC10, 9 on SSC11, 19 on SSC12, 35 on SSC13, 45 on SSC14, 29 on SSC15, 14 on SSC16, 11 on SSC17, 6 on SSC18 and 21 on porcine chromosomal X, respectively.
    1. Development of a livestock orthologous gene (LOG) database (Objective 3)

So far, we have collected full cDNA sequences of 33,308 human genes that are available at the NCBI Human Genome Resources. In our pilot study, these cDNA sequences were used as queries to search for the orthologous EST sequences in nine species: cattle, pig, chicken, horse, sheep, goat, rabbit, dog and cat. Computer programs were developed to execute this strategy en masse against the GenBank "est_others" database. The orthologous EST segments were collected based on criteria of a minimum length aligned by more than100 bp long with identify of more than 80%. Of the 33,308 human gene sequences studied, 18,867 were shown to have orthologous EST sequences in at least one of 9 livestock species described above, ranging from 1 to 249 EST hit(s) per gene. The numbers of genes with orthologous EST sequences found are 16,845 for cattle, 14,161 for pig, 6,547 for chicken, 4,203 for dog, 2,680 for horse, 1,134 for rabbit, 445 for goat, 203 for cat and 80 for sheep. Of 14,161 provisional porcine genes, 5935 have 1 – 2 EST hit(s), 3885 have 3 – 5, 2159 have 6 – 9, 1135 have 10 – 14, 541 have 15 – 20, 292 have 21 – 30, 193 have 31 – 50, and 21 have 51 – 106 tentative EST hits, respectively.

VI. Applications of Findings

  1. Bi-directional map-based chromosomal painting of the porcine and human genomes using both type I and type II markers will be useful in map-based positional cloning of candidate genes associated with QTLs as well as in map-based sequencing of the porcine genome.
  2. Selection of orthologous genes that are evenly distributed in the human genome at a 2 Mb interval as landmarks helps built comprehensive RH maps of the porcine genome.
  3. Development of a LOG database related to 33,308 human genes will play an important role in gene discovery, cDNA gap completion, gene functions and comparative genome mapping.

VII. General Project Plan (2003)
  1. Objective 1

    Experiment A. Update bi-directional map-based chromosomal painting between the porcine and human genomes using both type I and type II markers.

    Experiment B. Continue physical mapping of genes on SSC3 using a pig/hamster RH panel.

  2. Objective 2

    3. Experiment C. Select 300 microsatellite markers that are linked to functional gene regions as a panel for screening of QTLs for reproduction, growth and meat quality traits in pigs.

  3. Objective 3

    4. Experiment D. Continue LOG database development.

VIII. Publications

Jiang Z.H., O.J. Rottmann, J. Chen, H. Liu, O. Krebs and F. Pirchner (2002). A missence mutation in the follicle stimulating hormone receptor (FSHR) gene shows different allele effects on litter size in Chinese Erhualian and German Landrace pigs. J. Anim. Breed. Genet. 119: 335-341.

Jiang Z.H., J.S. Melville, H. Cao, S. Kumar, A. Filipski and A.M. Verrinder Gibbins (2002). Measuring conservation of contiguous sets of autosomal markers on bovine and porcine genomes in relation to the map of the human genome. Genome 45:769-776.

Jiang Z.H., H. He, N. Hamasima, H. Suzuki and A. M. Verrinder Gibbins (2002). Comparative mapping of Homo sapiens chromosome 4 (HSA4) and Sus scrofa chromosome 8 (SSC8) using orthologous genes representing different cytogenetic bands as landmarks. Genome 45:147 – 156.

Gibson J.P., Z.H. Jiang, J.A.B. Robinson, A.L. Archibald and C.S. Haley (2002). No detectable association of the ESR PvuII mutation with sow productivity in a Meishan x Large White F2 population. Animal Genetics 33:448-450.

King A.H., Z.H. Jiang, G.A. Rohrer, J.P. Gibson, D. Waddington and A.L. Archibald (2002). Use of radiation hybrid mapping to locate candidate genes for female reproductive traits on porcine chromosome 8. 28th ISAG meeting, Gottingen, Germany.

Mellvill J.S., Z.H. Jiang, A.M. Gibbins, J.A.B. Robinson and J.P. Gibson (2002). Chromosome-wide scanning of SSC3 for QTL associated with prolificacy in a Meishan x Large White population using a comparative mapping approach. 28th ISAG meeting, Gottingen, Germany.

X. Ye, J.A.B. Robinson, Z.H. Jiang, A.M. Gibbins and J.P. Gibson (2002). Genetic polymorphisms of histone deacetylase genes 1,3 and fatty acid binding protein 3, 4 and their effects on economic traits in pigs. 7th WCGALP meeting, Montpellier, France.

Melville J.S., A.M. Gibbins, J.A.B. Robinson, J.P. Gibson, A.L. Archibald, C.S. Haley and Z.H. Jiang (2002). Association of a single nucleotide polymorphism at the NCOA1 locus with prolificacy traits in a Meishan * Large White F2 population. 7th WCGALP meeting, Montpellier, France.

Jiang Z.H., A.B. Robinson, A.M. Verrinder Gibbins, J.P. Gibson, A.L. Archibald and C.S. Haley (2002). Mapping of QTLs for prolificacy traits on SSC8 using a candidate gene approach. 7th WCGALP meeting, Montpellier, France.

Jiang, Z. H., H.H. Cao, and J. J. Michal (2003). Piggyback on humans: a virtual integration of type I and type II markers mapped in the porcine genome. PAG Meeting XI. San Diego, USA.

Cao, H.H., J. S. Melville, Z.H. Jiang, J.A.B. Robinson and A.M. Gibbins (2003). A high resolution radiation hybrid map of porcine chromosome 6 relative to human chromosomes 1, 16, 16 and 19. PAG Meeting XI. San Diego, USA.