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Genome resource databases in horticutural crops

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  • 1. 1
  • 2. 1 P.Gangadhara Rao IARI Ph.D Vegetable Science GENOME RESORCE CONSERVATION OF HORTCULTURAL CROPS
  • 3. WHAT IS GENOME ? “All the genetic material in the chromosomes of a particular organism; its size is generally given as its total number of base pairs”. http://www.nbpgr.ernet.in:8080/repository/glossary.htm
  • 4. Types of genomic resources • Genomic, mitochondrial and chloroplast DNA • RNA (total, mRNA, short RNAs) • DNA markers, probes, primers • Vectors (cloning, expression, binary) • Cloned genes, mutated genes, promoters, reporter constructs • Libraries (sub-genomic; tissue, stage and treatment specific cDNA and EST) • BAC, YAC, PAC clone set from sequencing Projects Sequence information
  • 5. Genomic resources are employed Transgenic plants Cisgenic plants Molecular breeding Germplasm screening
  • 6. Application of Genomic Resources DETAILED UNDERSTANDING OF PLANT BIOLOGY  Plant architecture  Reproductive biology  Vernalisation  Photosynthesis and partitioning  Stress tolerance/resistance  Nutrient metabolism H.Sonah et al. 2011. Biotechnology Advances. 29:199–209.
  • 7. Utilisation of Genomic Resources CROP IMPROVEMENT  Markers: Molecular Breeding  Genes: Transgenic development  Allele mining and mutant generation  Assessing plant diversity  Comparative genomics  Understanding epigenomes  Genotyping H.Sonah et al. 2011. Biotechnology Advances. 29:199–209.
  • 8. Status of sequencing projects in horticultural crops up to August 2010 H.Sonah et al. 2011. Biotechnology Advances. 29:199–209.
  • 9. ESTs resources available in public database (A) vegetables, (B) fruits, (C) flowers, and (D) other miscellaneous horticultural crops up to August 2010. H.Sonah et al. 2011. Biotechnology Advances. 29:199–209.
  • 10. DNA BANKS Three kind of collections: 1.Total genomic DNA 2. DNA libraries 3.Individual cloned DNA fragments (probes, satellites etc.) Advantages: • Convenient experimental material • Easy to exchange and ready material for further manipulations Disadvantages: • In vitro regeneration (only cloned fragment replicated to precision) • Documentation • Require high level of skills for exchange and use • Problem of ownership and control of samples • Total genomic samples are non-renewable • Allow recovery of single gene not of genomes J.L.Karihaloo.2012.DNA Banking and international effects. Training on conservation on genomic resources.NBPGR.N.Delhi.
  • 11. Major world plant DNA banks 1. Australian Plant DNA Bank, Lismore, Australia (http://www.biobank.com) 2. DNA bank, Instituo de Pesquisas, Jardim Botanico de Rio de Janeiro, Brazil Brazil (http://www.jbrj.gov.br/pesquisa/div_molecular/bancodna/so bre_ing.htm ) 3. Missouri Botanical Garden, Missouri, USA (http://www.welbcenter.org/dna_banking.htm) 4. Royal Botanic Gardens, Kew, Surrey, Great Britain (http://www.kew.org/data/dnaBank/homepage.html) 5. South African National Biodiversity Institute DNA Bank, Kirstenbosch, South Africa (http://www.sanbi.org/frames/researchfram.htm ) 6. National Institute for Agrobiological Sciences, Japan (www.dna.affrc.jp )
  • 12. The Botanical Garden and Botanical Museum Berlin-Dahlem DNA Bank
  • 13. Microbiology Riken Bioresource Center DNA Bank; Animals: San Francisco Zoo DNA Bank, The Frozen Ark, The Ambrose Monell Cryo Collection, New York, The Animal Gene Storage Resource Centre of Australia, Conservation Genome Resource Bank for Korean Wildlife, Frozen Zoo, Austrian DNA Bank for Farm Animal Genetic Resources, National Plant, Fungi and Animal DNA Bank, Poland (Source: http://www.dnabank-network.org/Links.php ) Plant genomic resources banks Royal Botanic Gardens DNA Bank, Kew DNA Banking at the Missouri Botanical Garden The Australian Plant DNA Bank NIAS DNA Bank, Japan Bank at the Nationaal Herbarium Nederland DNA Bank Brazilian Flora Species Plant DNA Bank in Korea DNA Bank at Kirstenbosch, South Africa Trinity College DNA bank, Dublin
  • 14. Plant Genomic Resources Sequencing Mapping Microarray Proteomics Deciphering pathways
  • 15. DIFFERENT GENOMIC RESOURCE DATABASES • NCBI http://www.ncbi.nlm.nih.gov/ • SOL GENOMICS NETWORK http://solgenomics.net/ • BRASSICA GENOME GATEWAY http://brassica.nbi.ac.uk/ • CUCURBIT GENOMIC DATABASE http://cucumber.genomics.org.cn/page/cucumber/index.jsp • GRAMINACEAE TFDB http://gramineaetfdb.psc.riken.jp/
  • 16. CASE STUDIES
  • 17. NCBI is a good starting place • A very good and comprehensive site • http://www.ncbi.nlm.nih.gov/ • What is available there: – Databases – Tools – Education – Downloadable material
  • 18. NCBI website
  • 19. Finding a gene • Entrez – The underlying software that links the various resources and allows Boolean searches • BLAST – The Basic Local Alignment Search Tool (BLAST) – finds regions of local similarity between sequences by searching sequence databases and calculates the statistical significance of matches
  • 20. Using this information • Downloading the files – Various formats available • Finding Related genes/domains – What does your gene do? • Finding Taxonomic information – to select genes of interest for evolutionary comparisons • Finding Structural information – To identify significant regions of biochemical interest
  • 21. Ways to search • http://www.ncbi.nlm.nih.gov/
  • 22. The resources available are rich Many web-based and downloadable applications are available
  • 23. We can search many databases
  • 24. Searching all databases takes you to an Entrez links screen Clicking on one will take you to a subset of the data
  • 25. Protein files Clicking on a file will take you to a Genpept entry.
  • 26. GenBank file http://www.ncbi.nlm.nih.gov/protein/125924597?report=genbank&log$=prottop&blast_rank=1&RID=Y7NSNX8N016
  • 27. What can we do with the file? • You can reformat it using the Display button • You can download to various locations
  • 28. Displaying the file as a FASTA • FASTA is a useful format widely used by many bioinformatics software packages. • We can use it to conduct a BLAST Search to find sequences on the basis of homology
  • 29. Conducting a BLAST
  • 30. Types of BLAST • Basic BLAST – Search protein or nucleotide sequences against databases • Includes PsiBLAST, PhiBLAST, MEGABLAST etc – Search a combination using translated sequences • Blastx, tblastn, tblastx • Specialized BLAST – Specialized databases
  • 31. Using a protein file
  • 32. BLAST results
  • 33. Scrolling down reveals similar records
  • 34. B.V.Suresh et al., 2014.PLOS ONE: 9:(1) 86387.
  • 35. • OBJECTIVE mapping and a gene description • MATERIAL AND METHODS • S. lycopersicum Heinz 1706 reference genome sequence • Raw sequence reads of S. pimpinellfolium LA1589. • The RNAseq data was downloaded from short read archive of NCBI [accession number SRX118613 (leaf) SRX118614 (root) SRX118615 (flower) SRX118621 (mature green fruit)]. • QTL and EXPEN-2000 map coordinates were retrieved from SGN.
  • 36. The flow of the organization of the data in tomato genome database B.V.Suresh et al., 2014.PLOS ONE: 9:(1) 86387.
  • 37. Screenshots of a map view and a gene description page showing various features of tomato genome database. B.V.Suresh et al., 2014.PLOS ONE: 9:(1) 86387.
  • 38. FUTURE THRUST • Rapid development in molecular biology leaves behind all the allied crop sciences which are more applicable in crop improvement. • The gap between these crop sciences and genomics will widen with the changing scenario of technology. • These limitations present a number of challenges for technology development, data interpretation, and ultimately for integrating information from multiple disciplines. • Many economically important horticultural crops have very less or no. genomic resources available which need to be taken on priority basis. • One of the challenges in understanding genome structure of woody plants is unavailability of ESTs or other genomic resources. • Even after identification of trait specific genes from the genomic resources, their functional validation would be difficult