Agroecological impact of an in vitro biotechnology approach of embryo development and seed filling in legumes

Agron. Sustain. Dev. (2015) 35:535–552 DOI 10.1007/s13593-014-0276-8 REVIEW ARTICLE Agroecological impact of an in vitro biotechnology approach of embryo development and seed filling in legumes Sergio J. Ochatt Accepted: 21 November 2014 / Published online: 30 December 2014 # INRA and Springer-Verlag France 2014 Abstract Ongoing global climatic changes and growing demographic pressure have increased demand for agronomic resources and affected the agroecosystem by provoking a number of abiotic stresses that, added to biotic ones, result in physiological and metabolic disorders. Such stresses ultimately impact yield when it most needs to be improved, and understanding and resolving them is a major scientific and agronomic challenge of this century. However, many species are difficult to breed for stress resistance and improved yield for a number of reasons, ranging from a long life cycle (woody species), a reduced genetic background (most self-fertile, cleistogamous legumes) or conversely extensive heterozygosity resulting from an outbreeding nature, and also due to the mainly multigenic origin of such resistances. Biotechnologybased breeding would be an efficient alternative but, for recalcitrant crops, many attempts at in vitro regeneration met with varying degrees of success and often limited to a few genotypes, hampering exploitation of biotechnology approaches. To reduce the risk of undirected somaclonal variations amongst regenerants and transformants, it is better to produce them through somatic embryogenesis that recognises a single-cell origin but whose feasibility is also limited amongst species. There is also a need to fix the resulting genome once a novelty is obtained to ensure efficient heritability of improved traits acquired, which takes several generations in conventional breeding. Acceleration of generations through flowering and fruit set in vitro has been developed in various species including legumes. Haplo-diploidisation in vitro also offers a unique alternative to conventional methods, as it yields novel genetic combinations following doubling of haplotypes and regeneration of fertile plants S. J. Ochatt (*) INRA, UMR 1347 Agroécologie, BP 86510, 21065 Dijon Cedex, France e-mail: having gained homozygosity within a single generation. This review will examine the relationships between embryogenesis, stress and its impact on in vitro development of novel genotypes more apt for a sustainable agriculture. Keywords In vitro plant regeneration . Somatic embryogenesis . Gene transfer . Abiotic and biotic stress resistance . In vitro selection . Haplo-diploidisation . Genetic determinism . Embryo development . Seed filling . Legumes . Medicago truncatula . Flow cytometry . Endoreduplication . Auxin Table of Contents 1. Introduction: induced stress and embryogenesis in vitro go hand in hand 2. Legumes as a study subject 3. The model legume Medicago truncatula Gaertn 4. Seed development and the parallel with in vitro embryogenesis 5. Typical indicators of somatic embryogenesis in vitro 6. The study of seed (and embryo) filling 6.1 The central role of endoreduplication in the embryogenesis phase 6.2 The phase of morphogenesis 6.3 Auxin in embryo development, seed filling and endoreduplication 7. Functional validation of genes of interest by transformation 8. The link between cell cycle, embryogenesis and seed filling as affected by abiotic stress agents 9. Conclusion 10. References 536 1 Introduction: induced stress and embryogenesis in vitro go hand in hand Significant modifications have occurred in our planet over the last decades through an ever-increasing demographic pressure which encompassed much larger energy consumption and, in turn, is affecting the global climate (Branca et al. 2013). Demand for agronomic resources has hence substantially increased when most land apt for agriculture is already exploited and available land left is generally marginal. This has gravely affected the agroecosystem, and farmers are confronted with a number of abiotic stresses that add to those of biotic origin (e.g. pests and diseases), resulting in physiological and metabolic disorders that, ultimately, impact on yield when it most needs to be improved (Cuellar-Ortiz et al. 2008; Voisin et al. 2013; Yu et al. 2014). Therefore, understanding and resolving the impact of such stresses on yield is one of the major scientific and agronomic challenges of the twenty-first century. In this context, a number of biotechnology-based breeding approaches offer alternatives to conventional methods to generate novel genotypes with an enhanced resistance to both biotic and abiotic stresses and coupled with an improved yield (Ochatt et al. 2010; Rai et al. 2011; Pérez-Clemente and Gómez-Cadenas 2012; Campanelli et al. 2013). In many species, applying such approaches to preexisting genotypes for in vitro selection and/or gene transfer would significantly accelerate the breeding process, which would otherwise require a large number of successive generations to fix the novel resistance traits acquired in the genome thereby ensuring their heritability in the progeny. If coupled with the in vitro acceleration of generation cycles and/or with a faster genome fixation through haplo-diploidisation, breeding could be even faster. Care should be taken, though, to avoid regenerating non-true-to-type plants due to non-controlled and spontaneous somaclonal variations, as may arise through the regeneration of plants by organogenesis from calluses or explants instead of via a single-cell origin as in somatic embryogenesis-derived regeneration. Strategies have been developed for the induction of flowering in vitro in a number of species including legumes such as pea (Ochatt et al. 2002a, b; Ribalta et al. 2014), grasspea and barrel medic (Ochatt and Sangwan 2010) and lentil and faba bean (Mobini et al. 2014). Parallel to this, attempts have been undertaken at genetically manipulating flower induction for an increased precocity via the overexpression of genes such as Terminal Flower 1 (TFL1) gene in Arabidopsis thaliana (Hanano and Goto 2011), homologous of which have been found in many herbaceous species but also in fruit (Esumi et al. 2005, 2010; Wang and Pijut 2013) and forest tree species (Igasaki et al. 2008; Mohamed et al. 2010). On the other hand, regenerating haploid plants from unfertilised gametes followed by chromosome doubling of S. J. Ochatt regenerants to yield fertile double-haploid plants would permit to achieve homozygosity in a single generation (Lülsdorf et al. 2011; Pérez-Clemente and Gómez-Cadenas 2012). However, both pathways for haploid plant production (i.e. androgenesis from microspores and gynogenesis from unfertilised ovules) depend on the possibility to induce such gametes to undergo embryogenesis. Establishing reproducible and efficient strategies for the regeneration in vitro of fertile and true-to-type plants either issued from haplo-diploidisation as in chickp (...truncated)