title: Protocol for efficient regulation of in vitro morphogenesis in einkorn (Triticum monococcum L.), a recalcitrant diploid wheat species.
abstract: Einkorn (Triticum monococcum L.) is A-genome diploid wheat that has a potential to become a useful model for understanding the biology and genomics in Triticeae. Unfortunately, the application of modern technologies such as genetic engineering, RNAi-based gene silencing and genome editing is not available for einkorn as there is no efficient in vitro tissue culture and plant regeneration system. In the present study an efficient and simple protocol for plant regeneration via direct or indirect somatic embryogenesis and organogenesis has been developed. Various auxins used as sole inductors in einkorn displayed low effect for morphogenesis (0-8%) and plant regeneration (1-2 shoots per explant). The addition of Daminozide, the inhibitor of biosynthesis of gibberellins, together with auxin significantly improved the formation of morphogenic structures, especially when Dicamba (51.4%) and Picloram (56.6%) were used for combination; furthermore, the simultaneous addition of cytokinin into induction medium significantly promoted in vitro performance. Among the tested cytokinins, the urea-type substances, such as TDZ and CPPU were more effective than the adenine type ones, BA and Zeatin, for the regulation of morphogenesis; especially, TDZ was more effective than CPPU for shoot formation (11.73 vs. 7.04 per regenerating callus). The highest morphogenic response of 90.2% with the production of more than 10 shoots per initial explant was observed when 3.0 mg/L Dicamba, 50.0 mg/L Daminozide and 0.25 mg/L TDZ were combined together. Along with the identification of appropriate induction medium, the optimal developmental stage for einkorn was found as partially transparent immature embryo in size of around 1.0 mm. Although in the present study the critical balance between plant growth regulators was established for einkorn only, we assume that further the proposed strategy could be successfully applied to other recalcitrant cereal species and genotypes.
authors.affiliation: Institute of Basic Biological Problems RAS, Pushchino, Moscow Region, Russian Federation.|Branch of Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry RAS, Pushchino, Moscow Region, Russian Federation.
authors.forename: Dmitry
authors.initials: D
authors.lastname: Miroshnichenko
authors.affiliation: All-Russia Research Institute of Agricultural Biotechnology, Moscow, Russian Federation.
authors.forename: Inna
authors.initials: I
authors.lastname: Chaban
authors.affiliation: All-Russia Research Institute of Agricultural Biotechnology, Moscow, Russian Federation.
authors.forename: Mariya
authors.initials: M
authors.lastname: Chernobrovkina
authors.affiliation: Branch of Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry RAS, Pushchino, Moscow Region, Russian Federation.|All-Russia Research Institute of Agricultural Biotechnology, Moscow, Russian Federation.
authors.forename: Sergey
authors.initials: S
authors.lastname: Dolgov
mesh_terms: D003583:Cytokinins / Q000494:pharmacology; D004171:Diploidy; D007210:Indoleacetic Acids / Q000494:pharmacology; D009024:Morphogenesis / Q000187:drug effects / Q000235:genetics*; D063245:Plant Development / Q000187:drug effects / Q000235:genetics*; D010937:Plant Growth Regulators / Q000494:pharmacology; D014908:Triticum / Q000166:cytology / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development*
chemical_list: D003583:Cytokinins; D007210:Indoleacetic Acids; D010937:Plant Growth Regulators