Durable multigenic resistance for new apple varieties through cisgenesis

Transcription

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 Resistance is best for crop protection Appel scab: requires 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 Classical breeding for scab resistance in apple 1914: Vf-resistance discovered in crab apple Malus floribunda 821 in USA. World wide used source M.floribunda X Rome X Rome Beauty Beauty X ? X F ? X Resistant? X Resistant selection selection 80+ years later Vf-varieties with good quality Santana? X Resistant selection Elstar X Priscilla NL Santana

4 Scab resistance continued Seventh backcross generation Strong focus on fruit quality X Priscilla NL X Golden Delicious X Elise SQ- 159 SQ- 133 Natyra

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 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 M.micromalus extra source of resistance Origin of Resistance Cox's Dr.Oldenburg M.micromalus Elstar Vm Clivia Golden_Delicious Gala Vm Pinova Vm U family family 238, Vm

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 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 Cisgenesis - Transgenesis Cisgene (= gene from breeder s germplasm) - transgene (= foreign gene) Cisgenic transgenic Intragenesis: only native DNA. New combinations of promoter and coding region allowed. Cisgenesis is a strict case of intragenesis.

11 Cisgenesis: steps in the process Steps: 1. Isolate target gene from donor plant 2. 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 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 Micrografting cisgenic plantlet rootstock

14 Cisgenic shoot

15 Gala Santana SPVf2-1 SPVf2-2 SPVf2-11 SPVf2-15 LPVf2-1 LPVf2-4 LPVf2-16 RbcVf2-1 RbcVf2-2 RbcVf2-3 RbcVf2-10 RbcVf2-11 RbcVf2-12 Sporulation Sporulation on Vf2 transformants Sporulation (6 plants per event, leaf 1 to 4) 5 ef cdef cdef cdef ab abc ab ab a ab ab ab ab a a Transgenic lines

16 First cisgenic test orchard (Vf2) in Wageningen

17 First cisgenic test orchard (Vf2) in Wageningen

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 Ex: Multigenic resistance to late blight in potato

20 Graphical view of binary vector pbinplus: Rpi-blb3+Rpi-vnt1.1+Rpi-sto1

21 Multiple resistant transformants and control

22 Why cisgenesis? Pyramiding via cisgenesis is feasible. Classical breeding requires many generations and years, even with MAB Cisgenesis ~7 years One-step introgression without linkage drag

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 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) Cisgenesis is as safe as conventional breeding

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 support for transgenic and cisgenic apples From: George Gaskell et al., Europeans and biotechnology in 2010, Winds of change? A report to the European Commission s DG Research October 2010

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 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!! [email protected] [email protected]