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Chapter 19 Genomics and Agriculture

Chapter 19 Genomics and Agriculture . Applications of genomics approaches to agriculture. Contents. Background Agriculturally related sequencing projects Crop plant Farm animal Pathogens Genomics applied to trait improvement Breeding Transgenics, biotechnology and clones. Background.

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Chapter 19 Genomics and Agriculture

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  1. Chapter 19Genomics and Agriculture Applications of genomics approaches to agriculture © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  2. Contents • Background • Agriculturally related sequencing projects • Crop plant • Farm animal • Pathogens • Genomics applied to trait improvement • Breeding • Transgenics, biotechnology and clones © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  3. Background • Agriculture looks to genomics for the next “green revolution” (Gene revolution?) • Reasons: • POPULATION EXPLOSION (7 billion and 800 million malnourished) • Pace of traditional breeding is slow • Identify genes for useful traits • Relate a trait to genetic and physical maps and whole genome sequencing • Protect food chain and security (pathogens) © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  4. Genomics applied to agriculture • Approaches • Similar for crop plants and farm animals • Relating traits to genes • Relating genetic maps to physical maps • Rarely monogenic • QTL analysis (Quantitative Trait Loci) • DNA sequence • Problem of large genome size • So use syntenic relationships • ESTs • Genes can be manipulated, either through breeding or through genetic engineering, to remove deleterious traits and enhance desirable traits © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  5. Sequencing of plant genomes • Reference plant: Arabidopsis thaliana • No direct agricultural value • Related to cabbage and mustard • Reference for all plants • First plant genome sequenced (Dec. 2000) • Size: 130 Mbp • Number of genes: 28,000 • Segmental duplications • segmental duplications larger than 100 kb make up nearly 60% of the Arabidopsis genome • Evidence for past increase in ploidy • Only 35% of Arabidopsis genes are unique, while 38% belong to families of more than five members © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  6. Sequencing of crop-plant genomes • Reasons for sequencing rice first • Importance as crop • Largest food source for poor • Feeds half of world’s population (3 billion) • Demand likely to increase dramatically • 80% of daily calories in Asia come from rice • In Asia alone, demand will increase by at least 35%. © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  7. Rice genome • Smallest among grass genomes (Wheat, oat, rye, Barley, corn) • Few repetitive elements • Synteny with other grasses (recent evolution from a common ancestor approximately 50–70 million years ago) • Genetic and physical maps • Genomic resources • Over million ESTs • Efficient transformation © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  8. Efforts to sequence the rice genome International Rice Genome Sequencing Project (IRGSP) • Different efforts • Public: IRGSP: clone by clone/ • Beijing Genomics Institute: shotgun • Private: Monsanto/Syngenta shot gun • sufficient coverage to be able to map certain phenotypic traits such as plant size and fertility • Public performed 10x coverage • Two strains: Indica and Japonica • Gold standard for other cereal genomes • Microarray of rice can be used on maize RNA © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  9. Facts about the rice genome • Size: 430 Mbp (3.3 X Arabidopsis) • Number of genes: approximately 60,000 • Repetitive elements: Most in intergenic regions versus in introns in humans • Animals use alternative splicing and plants gene duplication? © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  10. Rice and Arabidopsis genomes Rice • No large areas of synteny • 80% of Arabidopsis genes have homologs in rice • Reverse not true • Only 50% of rice genes have homologs in Arabidopsis • 150–200 million years of divergence (Quick change) Arabidopsis © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  11. Genomics of other cereals • Maize: 3,000 Mbp • Wheat: 5,000 Mbp • Barley: 16,000 Mbp • Genome organization • Genic or gene-rich islands in the sea of retroposons © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  12. Sequencing strategies for grass genomes • Alternative approaches to genome sequencing • Methylation based: use non-methylated DNA by digesting genomic DNA with bacterial enzyme that destroys methylated DNA • Hybridization based: repetitive DNA fraction removed • EST collections based on synteny but private companies do not share the data © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  13. Genomics of other crop plants • Tomato, potato, soybean • EST collections • Woody species • Poplar and pine • Genome organization • No genic islands • Candidate-gene approach Because the genomes of most of the woody species are larger than even that of maize, genomic sequencing will probably have to wait until drastic reductions in sequencing costs arrive (How wrong was that prediction?). © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  14. Genomics of farm animals • Livestock farming = 30–40% of world agriculture • farm animals provide much of the protein in the daily diet in developed world • Poultry and livestock sales in United States > $70 billion • Disadvantages of genomics • Large sizes of farm-animal genomes like humans • Long gestation times • Difficulty of doing genetics • Nevertheless, genomics programs have been initiated for most of the major farm-animal groups, including pigs, cows, sheep, and poultry. © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  15. Genomics of farm mammals • Pig, cow, sheep • Draft sequencing • Compare to mouse, rat, and human • BAC libraries • Physical maps • EST libraries © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  16. Genomics of poultry • No reference genome • Different than humans • BAC libraries • > 300,000 ESTs • EST Microarrays • 22 K genes http://www.chick.umist.ac.uk http://www.genome.org/cgi/content/full/15/12/1692 © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  17. Sequencing of agricultural pathogens • Animal pathogens • Brucella suis (bacterium) • Infects animals, but can affect humans • Genome revealed to be similar function to plant pathogen Agrobacterium • Plant pathogens • Problem: large size of genomes • Agrobacterium, Phytophtora (potato blight), Fusarium bacterial pathogen that has invaded a cow’s milk-secreting cells, causing mastitis (inflammation of the mammal breast). © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  18. Bioterrorism issues: Food security • Intensive agriculture raises risks of disease spread • Example: outbreak of foot-and-mouth disease • Costed 48 billion to UK • Knowledge of pathogen genomes • Helps identify disease agent • Could be used in rapid-detection technologies • Could also be used a biological weapon by the US military who considered using Brucella in 1950s © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  19. Breeding • Domestication of plants and animals selected for valuable traits • e.g., temperament allowing bovines to be kept in captivity • Later, other traits selected for • e.g., milk production • Traits controlled by several genes • For each gene, different alleles © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  20. Quantitative traits • Major traits in continuous gradient (not yes and no like monogenic traits) • Controlled by QTLs • Infinitesimal model • Many genes, each with small effect • Major-gene model • A few genes, each with large effect • Genomic nature of QTLs: transcriptional control © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  21. QTL analysis • QTL analysis requires genetic and physical maps • Similar to association mapping in humans • Relate traits to markers • Or cross two subspecies with different traits • Both domesticated: cows • Wild plus domesticated boars but takes time to introgress © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  22. Genome scan for QTLs using crosses • Genome scan for QTLs in progeny: co-segregation • Relate trait to markers • Identifies interval on chromosome © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  23. QTL for tomato fruit size © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  24. Marker-assisted breeding • Once a QTL is found, it can be used to assist breeding • Even if the nature/function of gene is unknown • Markers on either side of the QTL can be followed during the breeding program • Introgress the QTL from one subspecies into another • Markers have to be very closely linked © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  25. QTL to gene • If genome sequenced: • Candidate genes in interval • If genome not sequenced: • Find syntenic region in sequenced genome • In QTL analysis of fat and lean chickens by scientists at the Roslin Institute, a QTL was identified that accounts for over 40% of the genetic variability in this trait. When this portion of the chicken genome was compared with the human genome, five genes that play a role in lipid metabolism were identified in the QTL interval. • To confirm identity: • Look for mutations • Microarrays and 2-D gels for expression analysis • Transfer gene and determine consequences © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  26. Genomics tools for breeding • Polymorphic markers • Microsatellites • SNPs • Expression approaches • Microarrays • 2-D gels • Bioinformatics • Databases © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  27. Improvement of plant traits • Stress resistance • Abiotic • Salinity • Drought • Biotic • Pathogens • Increased yield per acre • Decreased fertilizer utilization: negative effects on downstream waterways • Improved value-added traits: oil absorption by potato during frying or color of grains © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  28. Examples of plant breeding • Hybrid vigor • Corn with improved yield • Compare inbred parental lines with hybrids • Size of tomato fruit • Comparisons of wild relatives with crop plants and found a single gene for that © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  29. Improvement of animal traits • Growth rate • Meat quality • Disease resistance • Reproductive performance • Behavior © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  30. Example of animal breeding • Meat quality in pigs • Meat-to-fat ratio • Cross Chinese Meishan pigs with European Large White pigs • Meishan much fatter than European variety • Identified QTL for lean meat © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  31. Transgenic technologies • Goal: rapid modification of genes responsible for traits in plants and animals • Gain of function: • Overexpression • Ectopic expression • Loss of function: • Homologous recombination • Antisense or RNAi © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  32. Reverse-genetics example Gene that encodes muscle-specific transcription factor in mouse Myogenin required Homologous recombination used to delete gene Mice born, but can’tmake muscle targeting vector neo Tk genome locus myogenin selection neo product of homologous recombination selectable marker disrupts myogenin gene

  33. RNAi and antisense RNA Double-stranded RNA able to disrupt gene expression Cells have machinery that destroy double-stranded RNA: viruses/ cDNA Appears to be basis for the following: Interfering RNA (RNAi) Double-stranded RNA introduced into cells Antisense RNA Introduce complementary RNA Forms double-stranded RNA in cells

  34. Gene knockout techniques • Homologous recombination: low efficiency • RNAi leaky Deleted in KO AGL5 wt genomic AGL5 KO construct KanR agl5 KO genomic KanR © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  35. Transgenic plant technology Nopaline synthesis Tumor production • Agrobacterium-mediated transformation • Gene inserted into Ti plasmid • Agrobacterium cocultivated with plant • Ti plasmid transferred into plant genome • Selection with antibiotics T-DNA Nopaline utilization Ti plasmid T-DNA Transfer functions Origin of replication Agrobacterium gall on a cherry tree © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  36. Transformation of rice © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  37. Transgenic animal technologies • Introduction of genes directly into nucleus • Microinjection 1980s • Used to produce transgenic pigs, cattle, and sheep • Problems: inefficiency, random chromosomal insertion, tandem duplication © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  38. Nuclear transfer • The term “Nuclear transfer” or “cell nuclear replacement” better than “cloning” • Technique known since 1950s • Transfer of nucleus from adult cell to unfertilized egg with nucleus removed • Or fusion of adult cell with enucleated egg • Problem: abnormal development due to imprinting where one of the parental gene is shut off in embryogenesis • A cloned elite cow sold recently for over $40,000 © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  39. Animal cloning (1997) Scottish Blackface Finn Dorset Udder cell Egg cell Remove nucleus from egg cell Fuse cells with electric shock Fused cell grows into an embryo Embryo is placed in foster mother Cloned lamb is born Dolly Finn Dorset © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  40. Xenotransplantation (the transplantation of animal organs into humans) Pharmaceutical proteins • Alpha-1-antitrypsin • Applications in emphysema and cystic fibrosis • Problems in isolating from humans (contamination), yeast, or bacteria (processing) • Target in cell and then perform nuclear transfer • Produce as Milk protein and purify © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  41. Pet cloning A cat named Little Nicky, produced in 2004 by Genetic Savings & Clone for a north Texas woman for the fee of US$50,000 (CC or copy cat) A family dog Missy was cloned in 2008 (Bestfriends again) A dog named Booger for its Californian owner was cloned into 5 clones by SNL (Korea) 50K Goats, sheep, pigs, Cattle Horses? Cloned in 2006 for $150K Humans? Dinosaurs? Mammoths? Risks?

  42. Summary I • Need for genomics approaches in agriculture • Genomic sequencing • Crop plant • Rice • Farm animal • Livestock and poultry • Pathogens • Bioterrorism © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  43. Summary II • Genomics and breeding • QTLs • Traits • Transgenic technologies • Plant • Animal © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

  44. Questions • What is your opinion about genetic engineering of bacteria, plants, animals and human? • Write an essay and submit by November 18th 2011 © 2005 Prentice Hall Inc. / A Pearson Education Company / Upper Saddle River, New Jersey 07458

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