SABINE MEYRUEY1,LUCIE LE JEANNE1,MATHIEU THOMAS2,FRANCK-EMMANUEL LEPRETRE1,VERONIQUE CHABLE1 1 INRA, UR980 SAD-Paysage, 65 rue de Saint Brieuc, 35000 Rennes, France
2CNRS, Centre d’Ecologie Fonctionnelle et Evolutive, 1019 Route de Mende, 34000 Montpellier, France
Corresponding author : [email protected]
Key words: Fagopyrum esculentum, phenotypic and genetic diversity, in situ (on-farm) and ex situ conservation, participatory research
Summary: Few buckwheat varieties are available on seed market and many transformers, farmers and beekeepers are looking for local varieties. Buckwheat diversity is a major source for participatory plant breeding in order to improve agronomic performances, ecosystem services and quality of products. Phenotypic and genetic diversity were assessed within and among French buckwheat populations from three origins: on farm conservation, in situ conservation from French gene banks, and 2 available certified varieties. The first results show a high level of inbreeding, a little differentiation among the populations and a higher genetic diversity at intra-population level than at inter-population.Background
Buckwheat pancakes (“galettes”) are part of cultural and gastronomic heritage of Brittany (Bretagne, Western part of France). Nevertheless the major part of buckwheat transformed in France today is imported.
Buckwheat cultivation declined sharply in the XXth century but has recently known a renewed interest, particularly for organic and low-input farming. Buckwheat (Fagopyrum esculentum), a cross-pollinated Polygonaceae, has agronomic, ecological and nutritional qualities (attractive plant for pollinators, requiring few inputs and manure, with a short cycle, allelopathic effect, nutritional qualities) and various uses (production of flour, honey and hulled grains, gluten-free food or beer, cover crop, green manure, use of buckwheat hulls for upholestery filling…). However some bottlenecks limit development of buckwheat cultivation. The main factor is a strong year in, year out yield irregularity, but also a late harvest in automn which requires an efficient grains drying management, and crop contamination in fields by tartary buckwheat (Fagopyrim tataricum) which is very hard to sort out without specific material.
Selection and research on buckwheat are not carried out by public research nor by private companies in France. « La Harpe », selected by INRA in the 1960’s, is currently considered as the only marketed variety available in Brittany. This variety is the only one giving right to the « Blé Noir Tradition Bretagne » label, a PGI (protected geographical indication). Today, many farmers, beekeepers and transformers are willing to diversify local varieties of buckwheat.
Main chapter Introduction
Characterization of genetic diversity of buckwheat is necessary in order to initiate on farm breeding strategy. The objective is to increase agronomical performances and product quality mainly for low input and organic agricultures since very few varieties are available on seed market. Phenotypic and genetic diversity were assessed within and among French buckwheat populations from three origins: on farm conservation, in situ conservation from French gene banks, and few available certified varieties. The aims were to determine: i) the structure of genetic diversity within buckwheat populations according to their origin; ii) the genetic relationship among buckwheat populations and marketed varieties. Knowledge of diversity within and among buckwheat populations will help in the development of appropriate breeding strategies for participatory plant breeding and genetic resources (in situ and ex situ) management of French accessions.
Material and Methods
Molecular identification and diversity characterization was carried out on 22 samples (cf table 1).: 11 samples of peasant populations, collected on farm in 2013, 10 samples of accessions from INRA gene bank of Rennes and 2 samples of marketed varieties, “La Harpe” and “Drollet” which originated respectively from 2 French regions, Bretagne and Massif Central.
“La Harpe” ; “Drollet” Certified variety
“Brusvily” ; “Langourla” ; “Massif Central” ; “Petit gris de la Gacilly” ; “Saint Méen” ; “Saint Congard” ; “Saint Martin sur Oust” ; “Saint Nicolas du Tertre” ; “Limousin Faux de la Montagne”; “Montredon”
Population (French gene bank)
“Petit gris du Morvan” ; “Petit gris du Tarn” ; “Petit gris de Sidobre” ; “Petit gris de Bain de Bretagne”(JPC) ; “Petit gris des Hautes-Pyrénées” ; “Petit Prussien gris” (JPLR) ; “Petit Prussien” (AP) ; “Petit Prussien” (BM) ; “Petit Prussien” (JD) ; “Monédières” ; “Sarrasin Français”
Table 3: Origins of the buckwheat samples. The initial letters indicate the name of the farmer owning the population.
The samples were characterized during a trial experiment in summer 2013 in Le Rheu in Brittany. Each sample was sown manually. Ten plants per sample were described according to the list of descriptors currently used for buckwheat and established by IPGRI protocol (1994). Statistical analyses of phenotypic data were carried out with the software R : Analysis of variance (ANOVA) and Tuckey’s range test [HSD]).
The populations were genotyped with sixty-four genome-wide SSR markers showing polymorphism in literature (Ma, et al., 2009, Konishi, et al., 2006, Iwata, et al., 2005), using 15 plants per populations. Molecular analysis was performed after a DNA extraction from leaf samples using a standard protocol. The study of genetic diversity was carried out using GENETIX, Darwin and STRUCTURE softwares.
Results and discussion
These are preliminary results which have to be deeper analysed in order to confirm the observed trends.
Only three phenotypic quantitative descriptors were discriminating criteria for populations and varieties: number of primary branches per plant, number of internodes per plant and stem diameter. However, these descriptors did not allow a clear differentiation between all the populations neither the comparison between these populations and the variety “La Harpe”. Furthermore, a broad intra-variety diversity was observed.
Among the 4 on-farm populations named “Petit Prussien”, 2 appeared different from “la Harpe”according to the shape and the color of seeds and to some of the quantitative descriptors.“Petit Prussien” is indeed a local population some farmers and transformers are looking for because of its putative qualities. It can be hard to distinguish as some populations are wrongly thought to be “Petit Prussien” because of inbreedings.
Populations named “Petit Gris”, from on-farm and gene bank conservations, showed similar characteristics for seeds like the certified variety “La Harpe”. This result could be coherent with the putative origin of the variety “La Harpe” which was supposed to be bred from local Breton populations.
Only 9 out of 64 SSR markers were polymorphic. Two to 24 alleles were observed at these 9 loci. The level of polymorphism was studied according to allelic diversity so as to assess a first evaluation of genetic diversity within and among populations taking into account their origin (local/on farm populations or marketed varieties).
“La Harpe”, with an average number of 4.3 alleles per locus, ranged under the others. This low allelic diversity could be explained by the effect of selection, although the other marketed variety “Drollet” showed a high value of 6.1 alleles per locus.
Within population and among them, expected heterozygosity (mean = 0.62) is higher than observed heterozygosity (mean = 0.56). For some populations this heterozygote deficit could be explained by the small sample size during regeneration or seed multiplication (bottle neck effect) but as we also observe this deficit for certified varieties, which are cultivated in large fields, we suppose that the heterozygote deficit could be explained by a part of autogamy in buckwheat reproduction.
At a global level, populations showed a high level of inbreeding (average fixation index, FIS = 0,13) whatever the origin. The FST value (average Fst =
0,04) supported little differentiation among the populations. A higher genetic diversity was detected at intra-population level than at inter-population level whatever the origin of the samples. The large intra-population diversity is coherent with open-pollinated species.
Conclusion
Few microsatellite markers, from the panel established by several authors in the world, were polymorphytic for the French populations. This result indicated that only a small portion of the global genetic diversity of the species was represented in the French populations. Moreover, both results from molecular analysis and phenotypic descriptors revealed more diversity within population than among them whatever the origin (on farm, gene banks and on market). Such results suggested that French populations could be managed like a “crop metapopulation” (Louette, 2000). In respect to our breeding objective, the next step will be a comparison of French populations with accessions from other gene banks in the world to broader the working genetic bases. Then, performance and yield stability, pollinators attractiveness and quality will be assessed in order to study the relationship between these criteria and diversity level. Finally breeding strategies will be drawn to improve French populations.
Acknowledgements
This study is funding by Fondation de France, project "Sarrasin de pays", n°00038087. References
Iwata, Hiroyoshi, et al. 2005. Genetic diversity among Japanese indigenous common buckwheat (Fagopyrum esculentum) cultivars as determined from amplified fragment length polymorphism and simple sequence repeat markers and quantitative agronomic traits. Genome. 2005. Vol. 48, 3, pp. 367-377. 144. DOI: 10.1139/G04-121. Konishi, T, et al. 2006. Development and characterization of microsatellite markers for common buckwheat. Breeding Science. 2006. Vol. 56, pp. 277-285. 47. Louette, D, 2000. Genes in the field. On farm conservation of Crop Diversity, Brush S (ed), Lewis publisher 2000: Chapter 5, pp. 109-142.
Session B Poster B5