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.

Implementation of cell technologies has essentially improved the plant breeding process in agricultural crops in the world. The production of pure lines in cultivated crops, particularly among cross-pollinated species such as cucumber (Cucumis sativus L.) requires much time, labor and expense. Thus, the use of DH-plants for production of fully homozygous lines for one year becomes a very promising method for near cucumber breeding program. The major factor limiting the wide use of DH is a lack of effective protocol for large-scale plant production. In this review the historical facts with description of three main methods of DH-plant production were presented. By now these three methods have been such as parthenogenesis in situ induced by pollination with irradiated or chemically treated pollen; androgenesis in vitro including anther and isolated microspore cultivation in vitro; gynogenesis through ovule cultivation in vitro. Comparative analysis of published data with regard to the efficiency of the technology for DH-plant production was shown as well as advantages and limitations of each technology were described.

The anther derived androgenic plants of white head cabbage cv. Nadzeja and CMS samples were obtained. The cytological analysis of ploidy level of regenerated plants and doubled haploids obtained by treatment of meristems with colchicine was performed. On the basis of androgenic doubled hap loids of white head cabbage genotypes the seed plants were developed.

Relevance. In recent years vegetable crop Brassica rapa ssp. chinensis var. purpuraria (synonyms: Brassica campestris L. var.purpurea Bailey; Brassica rapa L. ssp. chinensis var. purpurea) is gaining popularity as an object of genetic and molecular researches, and as an economically valuable vegetable plant due to the high content of biologically active compounds and distinctive economically valuable traits. Effective technology for development DH-plants to accelerate the breeding process for this culture has not been developed yet, so research in this area is relevant.Materials and methods. The study included two varieties from the collection of Vavilov AllRussian Research Institute of Plant Industry (VIR): No. 1301 (China) and No. 1357(Netherlands). Both protocols standard unmodified and with addition of silver nitrate (AgNO3) in the medium for embryogenesis induction were used in experiments for production of DH plants from isolated microspore in vitro. Direct chromosome counting in meristem cells and flow cytometry were used to determine the ploidy of regenerating plants.Results. As a result of the study embryogenesis in B. purpuraria culture can develop with the use of a standard protocol as well as with the addition of silver nitrate that showed a positive effect on the induction of embryogenesis. The yield of embryoids varied depending on the genotype of the individual plant within the variety accession. The highest yield of embryoids was 40 embryoids/petri dish. The main problem at the stage of regeneration is that about half of the regenerating plants occurred to be albinos and were not viable. We show a high degree of spontaneous chromosome doubling in regenerated plants (all analyzed plants were doubled haploids). In total 38 regenerated plants were obtained from accession No. 1301. It was shown that four DH-plants had self-incompatibility after self-pollination, but seed progeny from other plants was obtained. The created material was taken for genetics study and breeding work.

The role of plant extracts in the process of induction of embryogenic structures and its following development in plants is revealed. Thus, the addition in the culture medium of the extracts diluted in DMSO increases the rate of gynoand androgenesis on 2.4% as compared with control. Experimentally proved that the plant extract derived from the reproductive organs of certain genotype increases the morphogenetic potential of isolated organs only of the same genotype.

The biotechnological methods of plant cell culture in vitro are represented in the article. Recent developments in the field of clonal micropropagation, androgenesis, gynogenesis, and genetic transformation of vegetable crops are shown. Importance of utilization of biotechnological approaches for successful development of agriculture is underlined.