1. www.gramene.org A comparative mapping resource for the grasses Gramene Workshop @ Plant Biology July 25, 2004 Molly Fogleman Doreen Ware Pankaj Jaiswal GRAMENE www.gramene.org

2. Topics General introduction to the grasses –Molly Fogleman Gramene: a community resource –Doreen Ware Answering biological questions with Gramene –Pankaj Jaiswal Open Discussion We invite you to give feedback on this workshop by completing our survey. Gramene poster # 902

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4. 49% of the world’s calories (human consumption) are provided by rice (23%), wheat (17%) and maize (9%) Wheat is the staple food for 35% of the world Rice is the staple food for almost half the world’s population Source: Evolution and Adaptation of Cereal Crops, 2002 Science Publishers, Inc. Cereals as a Food Staple

5. www.gramene.org Diets high in grains can lead to reductions in: Coronary Heart Disease Cancer Diabetes The USDA recommends 6-11 servings of grains/day The USDA Food Guide Pyramid Source: www.usda.gov and Whole Grain Foods in Health and Disease, 2002 American Association of Cereal Chemists, Inc.

6. www.gramene.org Sources: USDA and the National Agricultural Statistics Service's March 2004 Historical Track Records for United States Crop Production FAOSTATS May 2004 Note: Figures have been rounded USA and World Grass Acreage

7. www.gramene.org Photo Source: USDA - National Resources Conservation Service Amount of land farmed in the US and the World is decreasing due to: Urbanization Erosion High soil salinity levels Population vs. Acreage Agricultural Environmental Impacts in the US: 48% of the impaired river miles 41% of impaired lakes in the United States. Nutrient, sediment and pesticide runoff Source: EPA 2003

8. www.gramene.org Question: How to Feed a Growing Population? Genotypes (high yielding, pest resistance, drought tolerant, salt tolerant) from existing germplasms (gene pool)? Bioengineered Food? Photo Source: The Washington Post PhotoVoyage: Rice a Global Grain

9. www.gramene.org Source: National Plant Germplasm System (GRIN), July 2004

10. www.gramene.org Phylogeny of the Grasses Source: Kellogg, 1998, PNAS

11. www.gramene.org Macrosynteny: Markers, QTLs, and Genes are found in similar positions Source: MD Gale & KM Devos, 1998, PNAS

12. www.gramene.org Why use Rice as a leverage? –Smallest genome –Sequenced genome –Extensive genetic & physical map resources –Thousands of genes and quantitative trait loci mapped –Large mutant & germplasm collections

13. www.gramene.org Doreen Ware USDA-ARS Cold Spring Harbor Laboratory Gramene a community resource www.gramene.org

14. Genomics by Proxy candidate 1 candidate 2 candidate 3 Maize, Barley, Sorghum, Oat, Wheat… Rice trait

15. www.gramene.org Timeline and Funding Gramene www.gramene.org –Funded October 2001 –Superceded USDA RiceGenes –First release of the database January 2002 Funding Sources –USDA CREES IFAS –USDA ARS Specific Cooperative Agreement –NSF Research Coordination Network –NSF Plant Genome Initiative

16. www.gramene.org Web pages accessed by month

17. www.gramene.org Project Participants Community Collaborators –Publicly funded projects –Individual researchers Gramene Staff (Cornell and CSHL) –Curators Information content –Software developers Visualization tools and data management –Outreach Scientific community and Secondary Educators

18. www.gramene.org Collaborators and Contributors http://www.gramene.org/collaborators

19. www.gramene.org Community Resource Integrative set of web-based tools for discovery –Semi-automated –Curated Software applications –CMap: Generic Model Organism Systems Database Project (GMOD) www.gmod.org Controlled vocabularies –Open Biological Ontologies (OBO) Plant Ontology (POC workshop July 27 ) Gene Ontology Trait Ontology Environment Ontology

20. www.gramene.org What’s in Gramene High-throughput data –Rice genome –Rice proteins –Functional annotation of gene products –Grass EST collections Curated data –Genetic maps –Physical maps –Protein annotation –Mutant (phenotypic variant) –Quantitative Trait Loci (QTL) Documentation and Help Guide

21. www.gramene.org Search Tools Genome browser BLAST CMap viewer Marker Protein Ontology Mutants QTL Literature

22. www.gramene.org Web Interfaces Standard Customizable Links –Within the database –Between database –Data sources

23. www.gramene.org Navigation bar standard

24. www.gramene.org Genome browser display Data sets (Genbank, public projects, curated annotations) Semi Automated Alignment pipeline Ensembl browser for the database structure and visualization. Developed by EBI/Sanger for the human genome project

25. www.gramene.org Ensembl Genome Browser

26. www.gramene.org Pages provide links within and between databases

27. www.gramene.org Views are customizable Select tracks to display Compact and expand Color

28. www.gramene.org Genome annotation of rice Gene model –Validation –Comparative analysis using other cereal sequence –Tissue expression –Functional assignment (GO)

29. www.gramene.org Associate the biology of the organism with sequence Phenotypes: mutants and QTLs Integrated map feature genetic markers

30. www.gramene.org Comparative Map display Maps and Correspondences (public projects, curated) –Literature –Community curation –Alignment from sequence Database structure and browser –CMap –GMOD project

31. www.gramene.org Correspondences in comparative map display Rice Genome Assembly is the reference map in the comparative map browser Sequence features and hybridized markers provide the correspondence within species and between species maps

32. www.gramene.org Wheat 4D, Rice 3, and Maize 1

33. www.gramene.org …….add Sorghum C syntenic

34. www.gramene.org Protein Browser Data sets –SwissProt rice proteins, annotations and associated literature Ontologies (controlled vocabulary) Custom database structure and visualization software

35. www.gramene.org Protein Page

36. www.gramene.org KEGG

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38. SwissProt

39. www.gramene.org SwissProt linkback

40. www.gramene.org Waxy associations to the Ontology browser

41. www.gramene.org Links to search, features, literature

42. www.gramene.org Phenotypes assays What do you want to capture? How will you integrate this information into a database?

43. www.gramene.org Common fields for curation of mutants and QTL Observable/measurable difference resulting from: –one or more mutations (alleles) –at specific genetic loci (mapped genes) –that interact with the rest of the genetic background (germplasm accession) –under a given set of environmental conditions (temperature, light, moisture, nutrition, space) –as part of a biotic regime (i.e., exposed to specific microbes, insects, plants) –measured in given location(s) & time(s) (season, year) –at specific stage(s) of development.

44. www.gramene.org Ontologies used in Gramene Gene Ontology –Molecular Function –Biological process –Cellular location Plant Ontology –Anatomy –Development Trait Ontology

45. www.gramene.org What Ontologies Let You Ask Find all rice mutants in my favorite colinear region of rice associated with dwarfism. What genes within a starch content QTL are predicted to be involved in carbohydrate metabolism? Find protein orthologs between rice & maize whose stage-specific expression patterns have changed.

46. www.gramene.org QTL search with trait category “Development”

47. www.gramene.org Answering biological questions with Gramene Gramene workshop @ Plant Biology July 25, 2004 Pankaj Jaiswal

48. www.gramene.org What do we already know ? The rice disease resistance gene Pi-ta Genetically mapped to chromosome 12 Rybka et al. (1997). It has also been sequenced Bryan et al. (1997). Map a cloned rice resistance gene to its putative location in the rice genome Compare its position to that of other mapped resistance genes TASKS

49. www.gramene.org Download the amino acid sequence for the Pi-ta protein for rice from the NCBI website There are three protein accessions one of which is a hypothetical protein similar to Pi-ta. The other two, with accessions AAO45178 and AAK00132, are the Pi-ta sequences. (You may click on their respective links to verify that they are indeed the sequences of interest.)

50. www.gramene.org Copy the sequence Gramene BLAST Search

51. www.gramene.org BLAST results The following table will result. Choose the sequence with the highest score (GRMP = Gramene Peptide) Ensembl Protein Report page, which provides information about the identity, description, and structure of the protein. Gene Model id

52. www.gramene.org View Transcript and peptide structure Gene Model id TIGR gene Id export sequences Location of Pi-ta gene on the rice genome Gene report page Ensembl Gene Report page provides information about the identity, base pair location on the chromosome, description, and structure.

53. www.gramene.org Red window on chromosome indicates region shown in “Overview”. Red window on “Overview” indicates region shown in “Detailed View”. Putative Pi-ta gene on the rice genome Hover mouse to get popup menu with marker name. This is also the nearest marker to the Pi-ta gene Genome browser view

54. www.gramene.org Hover mouse over “repeat” feature to get the popup menu. Genes mapped to + strand. Genes mapped to − strand. Putative Pi-ta gene Gene neighborhood view

55. www.gramene.org bp view of the 100bp region The overview suggested that Pi-ta is found on BAC AL772421 The CMap links to the comparative map tool displaying the comparison of genetic Vs the physical maps Gene base pair view List of BAC clones representing the overview region of the genome.

56. www.gramene.org The middle map is the same as the genomic sequence map found in the Ensembl viewer. The latter two are genetic maps from the Japanese Rice Genome Project and Cornell University, respectively. To highlight additional features add more maps, go to the CMap comparative Map Viewer by clicking the N button on the sequence map Comparative map view

57. www.gramene.org Scroll down and click on map 12. CMap view of chromosome-12 assembly Allows a user to select its choice of maps, highlight and restrict the feature types used in building the correspondences for comparison

58. www.gramene.org Comparative view of sequence Vs genetic maps

59. www.gramene.org Using the two genetic maps, a cluster of five resistance genes map to the centromere region of chromosome 12. Pi-ta, which is known to reside on the genomic sequence AL772421, also maps to the centromere, as evidenced by the alignment of the pseudomolecule to the genetic maps.

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62. Mutant Page view Trait Ontology page view

63. www.gramene.org Protein page view

64. www.gramene.org Find sequence homologs !

65. www.gramene.org Find literature references !

66. www.gramene.org Search QTL for your favorite trait !

67. www.gramene.org Locate QTL on genetic map and draw comparisons to other maps !

68. www.gramene.org View the sequenced rice genome with features from multiple grass species aligned to the Genome Browser. Allows views of 2 megabases to 10 basepairs. Features support linking to internal pages and external reference resources. Find known rice proteins and learn about their functions. Provides external links to GenBank and SwissProt as well as internal links to the Ontology database and the Genome Browser. Search the Gramene database. Find rice genes with known phenotypes, such as this semidwarf-1 gene. Provides links to their mapped position in CMap. View maps from different grass species and construct comparisons between them. Find quantitative trait loci for multiple grass species. Provides links to their mapped position in CMap. A collection of useful links, including genomic projects, other grass databases and bioinformatic resources. www.gramene.org Useful Gramene website links, including links for Tutorials and Downloads. If you are new to our site, we suggest you review our tutorials before starting. Use the “Feedback” button for questions, suggestions and difficulties. We would like to hear from you.

69. www.gramene.org Site Map, Help Guides and tutorials

70. www.gramene.org Future plans Training and community outreach Provide information from other grasses on –Maps and markers –Phenotypes (mutant + QTL) –Genes and proteins –Maize genome sequences –Germplasm / genetic stocks Tools to study diversity in the grasses Raw datasets from genotype and phenotypes studies

71. www.gramene.org Gramene and you: a partnership Involve high school and college teachers to help generate interest in Biology (Dolan center-CSHL) Invite YOU (the authors) to curate your own dataset Your suggestions and feedback –Do you think we should organize an advanced level workshop ? –Suggestions on Gramene workshops in your institution ? –What new features would you like us to provide ? –Let us know how would you like to use the genomic and genetic information for your project. –Any other way we can help you ?