Convegno la mela nel mondo interpoma bz - 17-11-2012 1 - ton den nijs

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Convegno la mela nel mondo interpoma bz - 17-11-2012 1 - ton den nijs

  1. 1. Durable multigenic resistance for new apple varieties through cisgenesis Ton P.M. den Nijs and Henk J. Schouten, Wageningen UR Plant Breeding Bolzano (It), 17 November 2012
  2. 2. Resistance is best for crop protection Appel scab: requires 20-25 fungicide applications in North-West Europe Environmental and economical threat Breeding for resistance since over 60 years! Single source of resistance: Malus floribunda 821
  3. 3. Classical breeding for scab resistance in apple 1914: Vf-resistance discovered in crab apple M.floribunda 821 X X Rome Beauty M.floribunda 821 Rome Beauty Malus floribunda 821 in USA. 9433-2-2 X 9433-2-8 World wide used source ? X F2-26829-2-2 ? X ResistantResistant selection ? X selection 80+ years later Vf-varieties ? X Resistant selection with good quality Elstar X Priscilla NL Santana Santana
  4. 4. Scab resistance continued Seventh backcross generation 1972-010-033 X Priscilla NL Strong focus on fruit quality 1980-015-047 X Golden Delicious X Elise 1990-045-133 1990-022-159 SQ- 159 SQ- 133Natyra
  5. 5. Resistance management Crop protection evolves into gene protection We strongly advise to keep orchards with Vf- varieties clean One to two spraying at asco-spore release peak
  6. 6. R gene resistance breeding: durable protection? Many successes but also leading to “man-guided evolution” Multiple gene resistance presents higher barrier Pyramiding of different – if available- genes needed
  7. 7. M.micromalus extra source of resistance Origin of Resistance Coxs Coxs Dr.Oldenburg M.micromalus Elstar 238 - Vm Clivia Golden_Delicious Gala 1993-209-015 238 - Vm Pinova Pinova 1998-002-021 238 - Vm 2004-002-012 2004-002-012 U family U family 238, Vm
  8. 8. Pyramiding genes for resistance Combination breeding through cross-breeding and MAB (Molecular marker Assisted Breeding) Long-lasting process because of long juvenile period and linkage drag. Possible solutions:  Shorten juvenile period  Prevent linkage drag  CISGENESIS
  9. 9. Definition of Cisgenesis A cisgenic plant is genetically modified with one or more natural genes from a crossable donor plant. The gene is under control of its native promoter, contains its introns and terminator. No foreign genes, such as bacterial genes Same genes as in classical breeding No linkage drag
  10. 10. Cisgenesis - Transgenesis Cisgene (= gene from breeder’s germplasm) - transgene (= foreign gene) Cisgenic – transgenicIntragenesis: only native DNA. New combinations ofpromoter and coding region allowed.Cisgenesis is a strict case of intragenesis.
  11. 11. Cisgenesis: steps in the processSteps:1. Isolate target gene from donor plant2. Bring this gene into a high quality cultivar (genetic modification)3. Evaluate the cisgenic plants for trueness to type and target character, select out somaclonal variants
  12. 12. Cisgenesis in apple in practice We developed pMF1 for marker free transformation ETH (Zürich) isolated HcrVf2 We inserted it into susceptible cv. ‘Gala’ We micrografted the cisgenic plants on M9
  13. 13. Micrografting cisgenic plantlet rootstock
  14. 14. Cisgenic shoot
  15. 15. Sporulation on Vf2 transformants Sporulation (6 plants per event, leaf 1 to 4) 5 ef 4.5 cdef cdef cdef 4 3.5 Sporulation 3 2.5 abc 2 1.5 ab 1 ab ab ab ab ab ab 0.5 a a a 0 LPVf2-1 LPVf2-4 SPVf2-1 SPVf2-2 SPVf2-11 SPVf2-15 LPVf2-16 RbcVf2-1 RbcVf2-2 RbcVf2-3 RbcVf2-10 RbcVf2-11 RbcVf2-12 Gala Santana Transgenic lines
  16. 16. First cisgenic test orchard (Vf2) in Wageningen
  17. 17. First cisgenic test orchard (Vf2) in Wageningen
  18. 18. How to turn this into durable resistance ?Pyramiding different genes.We intend to pyramid three resistance genes Vf2 gene from Malus floribunda Additional resistance genes from apple (V25, Vr2) We isolated already Vr2. We are testing candidate gene for V25.
  19. 19. Ex: Multigenic resistance to late blight in potato
  20. 20. Graphical view of binary vector pBINPLUS: Rpi-blb3+Rpi-vnt1.1+Rpi-sto1
  21. 21. Multiple resistant transformants and control
  22. 22. Why cisgenesis?Pyramiding via cisgenesis is feasible.Classical breeding requires manygenerations and years, even with MABCisgenesis ~7 years One-step introgression without linkage drag
  23. 23. Why cisgenesis? High quality apple cultivar maintained Genetic make-up of the original cultivar is preserved. One or a few genes added. Especially important for outbreeding, vegetatively propagated plants (apple, potato, banana, grape, etc.)
  24. 24. Why cisgenesis?As safe as conventional breeding A cultivar with a ‘history of safe use’ is used. Only well known genes from classical breeding are added Escape of foreign genes via pollen flow to natural vegetation can be a problem for transgenesis. However, in case of cisgenesis the genes are taken from wild relatives. EFSA Journal 2012, (10) 2561. “Cisgenesis is as safe as conventional breeding”
  25. 25. Why cisgenesis? Consumers generally prefer cisgenesis to transgenesis ( Eurobarometer) Cisgenesis respects natural crossing borders Remains within the order of the creation “No strange genes in my food”.
  26. 26. support for transgenic and cisgenic apples From:George Gaskell et al.,Europeans and biotechnology in2010, Winds of change?A report to the EuropeanCommission’s DG ResearchOctober 2010
  27. 27. Conclusions Why cisgenesis? - Durable disease resistance due to pyramiding of R genes - Gain of time - Specific; only wanted alleles inserted - High-quality cultivar maintained - Preferred by consumers compared to transgenesis; natural genes from the crop species itself - Reduces genetic vulnerability due to single gene resistance
  28. 28. Acknowledgements Suxian Zhu (Wag UR Plant Breeding) for unpublished results on Phytophthora-potato Cesare Gessler’s group, ETH, Zürich Inovafruit, funding Thank you for your attention!! Henk.schouten@wur.nl Ton.dennijs@wur.nl

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