Effect of sowing dates of donor plants and concentration of 2,4-D on the purity of productive anther formation of common barley (Hordeum vulgare L.) in anther culture in vitro.

Creating dihaploid lines of agricultural plants is a labour-intensive but essential step in variety production in modern plant breeding. This stage allows significantly accelerate the process of creating new varieties of common barley and other crops. Barley digaploids are produced mainly by anther culture and microspore culture. The authors preferred anther culture in vitro. In the present study, the influence of climatic factors in the cultivation of donor plants on the yield of productive anthers at different sowing dates was established. The authors also identified the more stable culti- vars with a high anther production regardless of sowing date (Signal, Laureate and Eifel). Varieties showed the highest number of embryo-like structures formation at the first and third sowing dates (Zu Suren, Zu Zaza); and sorts with a high rate of productive anther formation at the second sowing date (Acha, Exploer) were identified. Different concentrations of 2,4-D in N6 medium on the frequency of embryogenesis and yield of productive anthers were studied. As a result of this study, the authors found that different concentrations of 2,4-D (1 mg/l and two mg/l) had no significant effect on the for- mation frequency of productive anthers in all the varieties studied. When the embryogenesis capacity of the cultivars was reviewed, all the samples were found to be positive in anther culture. However, the array Zu Suren had a significantly lower effective anthers yield than the samples Signal and Acha. As a result of correlation analysis, the authors found a close relationship between the length of the ear tube of donor plants and the frequency of formation of productive anthers (r = -0.69). A close relationship with the development of optimal microspore phase for the induction of androgenesis in anthers extracted from the ear tube with an average length of 6 cm was determined. This information can significantly speed up the selection of donor plants, but it is recommended to confirm the stage of microspore development microscopically for each new cultivar used.

Embryogenesis of European Radish (Raphanus sativus L. subsp. sativus convar. radicula) in Culture of Isolated Microspores In Vitro

The European radish is one of the most unresponsive crops in the Brassicaceae family to embryogenesis in in vitro microspore culture. The aim of this work was to study the process of embryogenesis of European radish and its biological features. In this study, the embryogenesis of European radish is described in detail with illustrative data for the first time. For the first time for the entire family Brassicaceae, the following were found: microspores with intact exines with ordered-like divisions; microspores completely free of exines; and a new scheme of suspensors attachment to the apical parts of embryoids. The morphology of double and triple twin embryoids was described, and new patterns of their attachment to each other were discovered. Uneven maturation of European radish embryoids at all stages of embryogenesis was noted. The period of embryoid maturation to the globular stage of development corresponded, in terms of time, to the culture of B. napus, and into the cotyledonary stage of development, maturation was faster and amounted to 17–23 days. The rate of embryoid development with and without suspensors was the same.

Androgenesis of Red Cabbage in Isolated Microspore Culture In Vitro

Red cabbage belongs to the economically important group of vegetable crops of the Brassicaceae family. A unique feature of this vegetable crop that distinguishes it from other members of the family is its unique biochemical composition characterized by high anthocyanin content, which gives it antioxidant properties. The production mainly uses F1 hybrids, which require constant parental lines, requiring 6–7 generations of inbreeding. Culture of isolated microspores in vitro is currently one of the promising methods for the accelerated production of pure lines with 100% homozygosity. The aim of this study is to investigate the factors and select optimal parameters for successful induction of red cabbage embryogenesis in isolated microspore culture in vitro and subsequent regeneration of DH plants. As a result of research, for the first time, it was possible to carry out the full cycle of obtaining DH plants of red cabbage from the induction of embryogenesis to their inclusion in the breeding process. The size of buds containing predominantly microspores at the late vacuolated stage and pollen at the early bi-cellular stage has to be selected individually for each genotype, because the embryoid yield will be determined by the interaction of these two factors. In the six samples studied, the maximum embryoid yield was obtained from buds 4.1–4.4 mm and 4.5–5.0 mm long, depending on the genotype. Cultivation of microspores was carried out on liquid NLN culture medium with 13% sucrose. The maximum number of embryoids (173.5 ± 7.5 pcs./Petri dish) was obtained on culture medium with pH 5.8 and heat shock at 32 °C for 48 h. Successful embryoid development and plant regeneration by direct germination from shoot apical meristem were achieved on MS culture medium with 2% sucrose and 0.7% agar, supplemented with 6-benzylaminopurine at a concentration of 1 mg/L. Analysis of the obtained regenerated plants, which successfully passed the stage of adaptation to ex vitro conditions by flow cytometry, showed that most of them were doubled haploids (up to 90.9%). A low number of seeds produced by self-fertilization in DH plants was observed.

Creation of an interspecific hybrid of Capsicum annuum L. and C. frutescens L. using biotechnological approaches

Relevance. Pepper is a common crop both for fresh consumption and for the preparation of spices. Recently, along with the increasing popularity of C. annuum L. pepper, there is increasing interest in other species of this genus, which have a number of breeding and important properties. The most important method of enriching the gene pool of cultivated plants is distant hybridization, through which valuable traits are transferred from wild species to cultivated ones. The development of a new variety is a lengthy process, stretching over several years. In this regard, breeders have faced the challenge of obtaining pure lines to create a pepper hybrid with desired properties by applying modern biotechnological methods that will accelerate this process. One of them is the method of microspore culture, which allows mass production of haploid plants, reducing the time for creating constant parental lines.Material and methods. The aim of the work was to create an interspecific hybrid of hot pepper (C. annuum L. x C. frutescens L.) with high ornamental properties, a complex of economically valuable traits, with good taste qualities. The research was carried out in the film greenhouse of FSBSI FSVC in the Moscow region. The research material was a variety population of hot pepper Capsicum frutescens Cz-544-14, used as a paternal line, which was heterogeneous, and a pure line of C. annuum L. (Pb-551) created by classical breeding.Results. The pepper hybrid F1 Christmas bouquet was created as a result of hybridization of species parental forms obtained by different methods (biotechnological and classical). To accelerate the production of an aligned paternal form of C. frutescens L., the technology of doubled haploids through microspore culture was used. As a result, doubled haploid plants meeting the planned model (compact low habit, purple fruit colouring in technical ripeness and red in biological ripeness) were obtained. The resulting hybrid combined all the necessary economic features: high ornamentality, compactness, bouquet arrangement of fruits, high taste and aroma. Thus, the use of remote interspecific hybridization in the breeding process in combination with biotechnological approaches can accelerate the production of new forms of hot peppers that meet the demands of the market.

Obtaining doubled haploids of Cucurbita pepo L.

Doubled haploids have been widely used worldwide in breeding programs and fundamental research as valuable homozygous material for about 100 years. The species Cucurbita pepo L. are represented by a huge variety of forms, include highly productive vegetable crops and have a wide distribution in the world. Despite the great economic importance, the creation of effective protocols to ensure stable production of doubled haploids in this species remains an urgent task. DH plants are of interest not only because of the acceleration of the breeding process, but also because of the realization of the huge potential of gametoclonal variability inherent in this highly polymorphic species. In this review, we analyzed the main technologies used for obtaining doubled haploids in vegetable crops of C. pepo: parthenogenesis in situ stimulated by treated/irradiated pollen, gynogenesis in vitro (unpollinated ovule culture in vitro) and androgenesis in vitro (anther/microspore culture in vitro). An analysis is presented of the research carried out from the beginning of the discovery of haploid plants to the current advances and evaluation of the prospects in the field of DH plant production. The main critical factors influencing the efficiency of each technology and its individual steps are considered. The developed technology of doubled haploids obtaining using non-pollinated ovary culture in vitro is presented. This technology allows to obtain up to 55 embryoids per one cultivated ovary (28 embryoids/ 100 cultivated ovules) To introduce haploid technologies into the breeding process it is necessary to evaluate the obtained plants for ploidy level. The use of direct counting of chromosomes in apical cells may present a certain difficulty in this species due to their large number (2n=40) and their small size. Depending on the level of laboratory equipment, ploidy determination using flow cytometry of cell nuclei and counting the number of chloroplasts in stomatal guard cells in the epidermis of the abaxial side of the leaf may be more convenient methods. The prospects for the use of molecular markers for assessment for homozygosity in DH technologies used, including C. pepo, are discussed in the review.

Doubled Haploids in Eggplant

Eggplant is a solanaceous crop cultivated worldwide for its edible fruit. Eggplant breeding programs are mainly aimed to the generation of F1 hybrids by crossing two highly homozygous, pure lines, which are traditionally obtained upon several self crossing generations, which is an expensive and time consuming process. Alternatively, fully homozygous, doubled haploid (DH) individuals can be induced from haploid cells of the germ line in a single generation. Several attempts have been made to develop protocols to produce eggplant DHs principally using anther culture and isolated microspore culture. Eggplant could be considered a moderately recalcitrant species in terms of ability for DH production. Anther culture stands nowadays as the most valuable technology to obtain eggplant DHs. However, the theoretical possibility of having plants regenerated from somatic tissues of the anther walls cannot be ruled out. For this reason, the use of isolated microspores is recommended when possible. This approach still has room for improvement, but it is largely genotype-dependent. In this review, we compile the most relevant advances made in DH production in eggplant, their application to breeding programs, and the future perspectives for the development of other, less genotype-dependent, DH technologies.

Results and prospects for the development of scientific research in FSBSI FSVC

The article reflects the issues of the current state of scientific research at the FSBSI FSVC, shows the directions and prospects for the development of selection and seed production of vegetable crops. The main mission of the selection and seed-growing work of the FSVC is the creation and reproduction of varieties and hybrids of vegetable and melon crops of a new generation, characterized by resistance to abiotic and biotic stressors, high productivity and product quality. One of the main ways to increase the efficiency of the breeding process is the use of modern methods of biotechnology and molecular genetics, which make it possible to obtain new genotypes and reduce the time of breeding. Success has been achieved in the creation of homozygous lines of cabbage, squash, cucumber, carrots. For the first time, it was possible to complete a full cycle of obtaining doubled radish haploids in microspore culture in vitro. The institution pays special attention to immunological research on the resistance of crops and new varieties to diseases, as well as protecting plants from pests and diseases. Phytosanitary monitoring, assessment and selection of resistant genotypes are carried every years. One of the important theoretical and practical problems solved at the institution is the development of environmentally friendly products based on preparations obtained from plants with a high content of biologically active compounds with adaptogenic properties. In the center, a new direction of research in traditional breeding has received significant development - the creation of varieties of vegetable plants with an increased content of biologically active substances and antioxidants, on the basis of which functional products are created. The persistence of modern varieties and hybrids of table carrots, depending on the biochemical composition, has been studied. Methods for enriching vegetable plants with micronutrients are being developed. A large amount of scientific research is being carried out on the development of technologies for the cultivation of new varieties and hybrids, as well as technologies for their seed production. At the branches located in different soil and climatic conditions, hospitals were established to study the interaction of factors controlling soil fertility and the productivity of vegetable crops in agrocenoses (crop rotation, soil treatment systems, fertilizers and plant protection). In general, at the FSBSI FSVC, all research is aimed at developing and improving methods in breeding and seed production of vegetable crops, creating new breeding achievements and developing varietal technologies for their cultivation.