Annual Report of Multistate Research Activity

PROJECT NUMBER: NRSP-8

PROJECT TITLE: NRSP-8: Swine Species Genome Committee

PERIOD COVERED: January 1 to December 31, 2002

DATE OF THIS REPORT: February1, 2003

ANNUAL MEETING DATE(S): January 11, 2003

PARTICIPANTS: (names in bold represented Stations or administrative advisors)

Participant

Institution

Participant

Institution

Aaron Singsey

Biogenetic Services, Inc.

Shuhong Zhao

Iowa State Univ.

Akiko Takasuga

Sirakawa Inst. Of Animal Genetics

Stacey Meyers

Univ. of Illinois

Ana-Maria Gaseneau

Iowa State Univ.

Tim Smith

USDA/ARS MARC

Archie Clutter

Monsanto

Tom Rathje

Danbred North America

Atabak Royaee

USDA/ARS BARC

Tun-Ping Yu

Sygen International

Audra Kazlauskas

Univ. of Illinois

Udaya Desilva

Oklahoma State Univ.

Bart Tungerius

Wageningen University

Ulaus Olek

Univ. Bonn Biopsytisc

Brandy Marron

Univ. of Illinois

Zhihua Jiang

Washington State Univ.

Cathy Ernst

Michigan State Univ.

Zhiliang Hu

Iowa State Univ.

Chad Bierman

Babcock Genetics Inc.

Charles Otieno

Iowa State Univ.

Chris Bidwell

Purdue Univ.

Chris Tuggle

Iowa State Univ.

Christian Looft

Univ. of Kiel

Craig Beattie

Univ. of Nevada

Dan Nonneman

USDA/ARS MARC

Daniel Pomp

Univ. of Nebraska

Dave Burt

Roslin Institute

Deb Hamernik

USDA-CSREES

Diane Moody

Purdue University

Gary Rohrer

USDA/ARS MARC

Glenn Zhang

Oklahoma State Univ.

Honghe Coi

Univ. of Guelph

Jack Dekkers

Iowa State Univ.

Joan Lunney

USDA/ARS BARC

John Byatt

Monsanto

John McEwan

AgResearch

Jun Heon Lee

Chingnam Nat’l Univ., Korea

Kelly Swanson

Univ. of Illinois

Larry Schook

Univ. of Illinois

Lauree Rind

Univ. of Illinois

Lee Alexander

Univ. of Minnesota

Margaret OKomo-Adhiambo

Univ. of Nevada

Mark Thallman

USDA/ARS MARC

Matt Ehrhardt

Univ. of Illinois

Max Rothschild

Iowa State Univ.

Michael Grosz

Monsanto

Micheal Murtaugh

Univ. of Minnesota

Rick Van Wyle

Inst. Of Pig Genetics, Netherlands

Serguei Golovan

Univ. Guelph

BRIEF SUMMARY OF MINUTES OF ANNUAL MEETING:

Invited Speakers presented their talks:

"Molecular dissection of an imprinted QTL on SSC2 with major effect on muscle mass"

"Using cDNA microarrays to study ovarian follicle development in pigs selected for

increased ovulation rate"

"Bioinformatics update"

"Use of Real-time assays of immune gene expression to assess genetic basis of disease resistance"

Experiment Station Reports – see h ttp://www.genome.iastate.edu/community/NRSP8/2002_index.html

Indiana – Diane Moody

Iowa – Max Rothschild, Chris Tuggle

Michigan – Cathy Ernst

Minnesota – Lee Alexander, Mike Murtaugh

Nebraska – Daniel Pomp

Nevada – Craig Beattie

USDA/ARS BARC – Joan Lunney

USDA/ARS MARC – Gary Rohrer

Washington – Zhihua Jiang

Illinois

Kansas

North Carolina

Illinois

USDA, MARC

Swine Genome Coordinator’s report – Max Rothschild –see: http://www.genome.iastate.edu/community/NRSP8/2003_2002_index.html

Administrative Advisors’ report

Discussion Items

NRSP-8 Swine Committee Business Meeting, Max Rothschild, chair

New Business Items:

1) Location of NRSP-8 Swine Species Genome Committee for 2004

Daniel Pomp moved that the 2004 meeting be held at the Plant and Animal Genome Conference in San Diego, CA, as a joint meeting with NC-1004.

Joan Lunney seconded the motion.

Motion passed.

2) Industry Representative

Dr. Tom Rathje, Danbred North America, was introduced as the official industry representative to the NRSP-8 Swine Species Genome Committee.

3) Electronic reports

Max Rothschild reminded participants that electronic copies of station reports are needed by the end of the week.

4) Officers

Diane Moody will serve as the committee chair for the upcoming year.

Joan Lunney volunteered to serve as secretary.

Key Discussions:

1) NC1004 Update. Daniel Pomp announced that the first NC-1004 meeting will be held May 29-31 in Hastings and Clay Center, Nebraska. Brad Freking will serve as the local host.

2) Development of Pig Microarrays. Max Rothschild reported that he has begun investigating options of using Swine Genome Coordinator funds to develop and purchase a general pig microarray. He reported initial discussions with Agilent and Affymetrix had not led to feasible agreements because of intellectual property and cost issues, respectively. Additional options, including spotting cDNA collections at Michigan State University, or synthesis of long oligonucleotides through a commercial source such as Qiagen, were presented. Considerable concerns regarding quality control issues with the cDNA spotting approach were expressed. It was suggested that validation and curation of cDNA clones from multiple sources would be highly problematic. However, Michigan State has experience with quality control issues and is willing to make individual clones available to researchers for validation of specific array elements. Concerns relating to the synthesis of oligos included the ability to design suitable oligos for 10,000 sequences given the current publicly available sequences. Questions regarding the bioinformatics that would be used to select sequences and design oligos were also raised. Representatives from Qiagen addressed many of these questions and provided information describing their bioinformatics tools. It was decided that a committee would make a final recommendation to the Genome Coordinator regarding which approach to pursue, and that plans to make an initial pig microarray would move forward in the coming months.

Assigned Responsibilities/Deadlines/Target Dates:

A committee was established to review options for making a pig microarray and provide a recommendation to Max Rothschild by February 1, 2003. Committee members include:

Chris Tuggle, co-chair Cathy Ernst

Daniel Pomp, co-chair Diane Moody, Mike Murtaugh

 

ACCOMPLISHMENTS AND IMPACTS:

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

Production of high quality comparative maps requires the addition of genes to the map with which to compare with other species. This year a number of stations (IA, IN, MI, NE, NC, NV, USDA-BARC, USDA-MARC, WA) contributed to the comparative map. Examples of this mapping includes mapping of candidate gene loci (IA, NC, USDA-BARC), SNPs from STS (MI, USDA-MARC) and extensive mapping of genes using EST information (IA, IN, MI, NE, NV, WA, USDA-MARC). It is likely that the comparative map made more progress this year compared to any other previous year in that over 3,000 gene were added to the comparative map. These advances are likely to continue given that there is a new 12,000 rad RH panel and other tools.

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

Several experiment stations contributed new anonymous markers to the pig RH and genetic linkage maps. The MN station is involved in a large project to identify new microsatellites. They have 1,770 primer pairs developed and have mapped 771 new microsatellite markers on the IMpRH panel. These new markers have been published and will be useful for other stations. The USDA-MARC station has been developing SNPs and is creating a SNP map for future association work. Several stations (IA, MI, NE, NV, NC) are mapping new gene markers using ESTs and candidate genes. In total, more than 3,000 markers of this type have been added to the physical map. Other stations (IA, NE, IN, USDA-MARC, WA) are using information in the human to predict mapping location in the pig.

USDA-MARC is fine mapping the region of a Quantitative Trait Locus (QTL) for traits affecting litter size on SSC8. The development of 3 new resource populations for the identification of growth, meat quality and carcass merit QTL has been reported by the IL, IA and MI stations. A Berkshire x Duroc F2 population is under development in IL and a Duroc x Pietrain F2 population is under development in MI. The MI family is progressing nicely and over 500 F2 animals have been born and phenotyped. A Berkshire x Yorkshire F2 population at the IA station has been completed and 40 traits were measured on 515 animals. A primary and genome scan using 125 microsatellite markers and a secondary scan using 165 markers has revealed over 100 putative QTL significant at the chromosome wide level and 20 of these were significant at the genome wide level. In addition, many imprinted QTL have been identified. The underlying genes for at least two of these QTL have been identified. Several stations are advancing beyond QTL analysis and doing functional analysis. A number of stations (IA, IL, IN, MI, NE, NV, MN, USDA-MARC, USDA-BARC) are examining differential expression using arrays or real time PCR techniques. Such work will suggest underlying genes possibly responsible for QTL.

Progress Toward Objective 3: Expand and enhance internationally shared species genome databases and provide other common resources that facilitate genome mapping.

Database development has continued to be enhanced by efforts by the US Pig Genome Coordinator. A central web site exists (www.genome.iastate.edu) that contains data information on a variety of items related to pig gene mapping. This includes information of linkage and physical maps, tools to finds sequence information and reference materials. Several stations (IA, IN, MI, NV, WA) are developing their own databases to catalogue information generated by their EST or other expressions projects. At IA there is a database with nearly 100,000 ESTs. Two stations (MI, NV) are developing their own EST databases and will combine them with expression data. At Purdue (IN) particular emphasis has been placed on the ability to integrate different types of data, such as phenotypic, gene expression, and SAGE data, with other efforts focused on mapping and annotation of pig gene sequences. A livestock orthologous gene (LOG) database is being developed (WA) to combine BLAST searches against GenBank databases using human cDNA sequences as queries, identification of orthologous gene/EST segments in livestock species, and generation of tentative or orthologous consensus sequences by assembling and ortholog assignment and comparative mapping. Efforts to combine these databases are underway.

An effort to share resources continues. BAC libraries and filters were supplied by the US Pig Genome Coordinator to 12 labs in the US. Tissue specific EST libraries have been constructed at several stations (IL, IA, MI, MN, NE, NV, USDA-MARC). Tissues represented by these libraries include skeletal muscle, placenta, anterior pituitary, ovary, conceptus, fetus, hypothalmas, alveolar macrophages, peripheral blood cells, spleen, thymus, lymph node and bone marrow. Also one station (IN) has been developing libraries for SAGE analysis. Efforts to produce a porcine oligo array using these ESTs are underway. Slides from this array will be shared with the porcine community using coordinator funds.

Impacts:

Work done by the Swine Genome Committee of NRSP8 has significantly enhanced the knowledge of the pig genome. The mapping of additional genes and ESTs facilitates the usefulness of the recently published human genome sequence because maps comparing the location of genes on human chromosomes with their location on pig chromosomes provide a rich source of candidate genes for QTL. A significant number of genes (ESTs) were added primarily to the pig physical map. These new gene markers will aid in the positional cloning of genes at QTL because having many gene markers available in regions harboring QTL considerably speeds the process of determining the exact position of the QTL and leads to its eventual identification. Toward this end genes continue to be identified for traits such as growth rate, backfat and meat quality. The most notable being CAST which affects tenderness. As more genes at QTL are identified, it will be possible to improve the efficiency and reduce the costs of pork production by improving the speed and accuracy of selection programs. The development of EST libraries has increased and this has allowed for the functional study of many genes simultaneously. Work is progressing to develop pig gene arrays that will help researchers understand the functional biology of gene action. This array technology, combined with bioinformatics and new genome database development has the opportunity to speed identification of specific genes having a major effect on growth, carcass merit, reproductive efficiency and health and promises to facilitate the production of a significantly improved pork product at reduced costs. American agriculture will thus be more competitive on the world market and the consumer will ultimately benefit.

Plans for the coming year:

Work planned by the Swine Genome Committee for next year includes further refinement of the pig gene maps. New anonymous markers will be identified and located on the genetic linkage map. Several stations are planning to sequence and map a significant number of ESTs that will aid the physical and comparative maps. New resource populations are under development and QTL identification will accelerate. Fine mapping of regions containing previously identified QTL will continue and identification of useful candidate genes will continue. Database development will continue and be directed at improving tools for genome researchers.

PUBLICATIONS Published in 2002 from reporting stations:

Bertani GR, C.D. Gladney, R.K. Johnson and D.