Ecological and biological bases of creating source material of sea buckthorn ( Hippophae rhamnoides L.) on adaptability and productivity for further breeding

. The research relevance is determined by the constant striving to improve and enhance sea buckthorn genotypes by using different genetic resources and breeding methods to achieve better adaptability, higher productivity, and fruit quality. The research aims to expand the genetic diversity of sea buckthorn, which combines high productivity and quality for further breeding. Phenology, morphological studies, and evaluation of breeding material for economically valuable traits were carried out


INTRODUCTION
The development of new genotypes of sea buckthorn with high adaptability to changing climatic conditions, high productivity and exquisite fruit quality is an extremely important task to meet the needs of the population in food resources.Resistant to adverse biotic and abiotic environmental factors, highly productive varieties of agrobioproducts fully solve the problems of energy and resource conservation, protection of the biosphere from pollution by pollutants and management of sustainable development of agroecosystems.In works (Moskalets et al., 2020;Todd, 2022), it is described that sea buckthorn is dioecious (staminate flower formula: *K (2) C 0 A 4 G 0 , pistillate flower -*K (2) C 0 A 0 G 1 ), is a diploid (2n=24), wind-pollinated, facultative parthenogenesis, perennial shrub or tree with thorny or nonthorny branches, an ancient crop with modern significance and is important among the cohort of rarely cultivated fruit and berry plants, as its fruits are a source of biologically active substances.
Z. Ciesarová et al. (2020) pointed out that the autochthonous populations of Hippophaer hamnoides L. are more widespread in Eurasia than other species and, depending on the physical and geographical area and place of growth, plants of this species can be in the form of a bush (mostly in the Western European part, up to 10 m high) or a tree (Sichuan, China; Åland Islands, archipelago in the Baltic region) up to 10-18 m high.The average life span of sea buckthorn plants is 30-40 years, with a maximum of 80 years.Studies (Husain et al., 2018) indicate that sea buckthorn was favoured by climate, soil, topography, and other environmental factors that led to the formation of a rich germplasm resource in the ecosystems of China, India, Pakistan, Siberia, Germany, Denmark, Romania, Hungary, Tajikistan, etc.However, the agronomic potential of many Hippophae species remains underutilised or undiscovered.J. Zhao et al. (2023) described that sea buckthorn in Ukraine is a source of vitamins, an ingredient in food combinatorics for the production of healthy foods and can take an important place in the structure of an orchard.According to Serhii Ostapets, CFO of Sady Donbasu (2022), producers should stop creating new apple orchards and start focusing on other crops, including sea buckthorn.
The authors (Stobdan et al., 2022) noted that the high level of sea buckthorn diversity offers excellent opportunities for the genetic improvement of plants.The main objectives of most sea buckthorn breeding programmes are yield, fruit size, thornlessness, fruit size, oil content, ripening time, frost resistance, resistance to desiccation disease, fruit taste, pollen, and ornamental value.The first sea buckthorn breeding programmes began with massive selection from natural populations.This method is still an established practice, but it is gradually being replaced by mutation and hybridisation.
Studies of genetic resources of the genus Avena sativa (Bartish & Thakur, 2022) have accumulated valuable information on the history of evolution, biogeography, genetic diversity in populations, population structure, and genes with putative specific adaptive functions in different species and taxa.Attempts to disseminate foreign varieties of sea buckthorn in production have been unsuccessful due to their low adaptability to significant fluctuations in weather conditions throughout the year (Höhn et al., 2019).First of all, plants of foreign varieties cannot withstand low temperatures in winter, are not sufficiently drought-resistant, are affected by pathogens and specific pests, and are characterised by low productivity and fruit quality.According to L. Dienaite et al. (2021), the low level of genetic variation in sea buckthorn populations requires work to develop source material of different ecological and geographical origins.In China, the main emphasis was placed on local forms, given their high adaptability and wide ecological valence, and introduced varieties that were used in hybridisation with local ones.
The use of the diversity of natural and grassland ecosystems in the breeding process, given the narrowing of sea buckthorn variability, will expand its genetic base for further breeding for adaptability, productivity, and quality to overcome food problems, including providing the processing industry with healthy raw materials for the production of products for healthy eating.The research aims to develop new forms of sea buckthorn that combine high adaptability and productivity for further breeding.

MATERIALS AND METHODS
The research was carried out during 2017-2022 at the Institute of Horticulture of the National Academy of Agrarian Sciences of Ukraine (NAAS).Based on the designed model of the variety, a programme was developed, which consistently substantiated the ways to create new sea buckthorn source material.The first stage of research was carried out in 2015-2016, which involved studying the populations of fallow land in the Polissia and Forest-Steppe zones and selecting samples with desirable economic traits.Locations of sea buckthorn fallow land populations were examined and analysed according to the methodology of S. Tkachyk et al. (2016).The second stage of the breeding involved the introduction of selected sea buckthorn forms into the hospital of the Institute of Horticulture of the National Academy of Agrarian Sciences of Ukraine for further study and involvement of the best ones in further breeding.The soil type of the site is dark grey podzolised, medium loamy on loess loam, characterised by the following agrochemical features: humus content -2.3%, easily hydrolysable nitrogen -8.72 mg/100 g of soil, mobile phosphorus, and exchangeable potassium -27.5 and 24.8 mg/100 g of soil, respectively, pH -6.8 (Puzniak et al., 2022).Planting scheme of sea buckthorn plants 4×2 m and 4×1.5 m.
Before the polymerase chain reaction, it was necessary to defrost, mix and centrifuge the special solution to the main components of the polymerase chain reaction (except for the DNA polymerase enzyme involved in DNA replication) and total DNA preparations.The process of DNA fragment fractionation involved the use of the following reagents: UltraClean TM agarose (MO-BIO); LB buffer (10 mM LiOH, H 3 BO 3 to pH 8.5); ethidium bromide 10 mg/ml concentration; GeneRuler TM DNA LadderMix molecular weight marker (ThermoScientific), distilled water, total DNA preparations.
An agarose gel was produced at a concentration of 1.2% in 1×LB buffer by weighing the required amount of agarose and placing it in a Simax thermobarrel to the desired volume of buffer.The mixture of agarose and buffer was then brought to complete dissolution in a microwave oven.Next, the heat-resistant jar was cooled to 60°C under a constant stream of tap water, while stirring the mixture continuously.Ethidium bromide was used as a dye at a concentration of 0.50 µg/ ml.The plates were sealed with rubber spacers and the combs were placed on the required number of wells.The plates were filled, and the agarose was allowed to solidify for 30-60 min under a fume hood.Before inserting the plate into the chamber, the combs were removed.The gel plate was placed in the chamber so that it was completely covered with the buffer solution.In each well, 8 µl of sample (for PCR products) or 5 µl (for total DNA) and 1 µl of 6× application buffer was carefully added.
Next, the GeneRuler TM DNALadderMix molecular weight marker (ThermoScientific) was used (plycon sizes, bp: 100, 200, 300, 400, 500, 600, 700, 800, 900, 1000, 1200, 1500, 2000, 2500, 3000, 3500, 4000, 5000, 6000, 8000, 10000) to determine the size of the amplification products.For this purpose, the electrode voltage was set to 3 to 7 V/cm, depending on the size of the desired amplicons.The electrophoresis time is 2 to 4 hours.To obtain electrophoregrams, UV light was used in conjunction with a Canon camera and the GIMP graphic editor.The choice of DNA markers and primer design and their characterisation of sea buckthorn SSR primer marker loci are presented in Table 1.

Source: compiled by the authors
Total DNA was extracted from the plant material by the RT-PCR method (cetyltrimethylammonium bromide).Then, 0.030-0.040g of the sample was taken and placed in a 1.5 ml Eppendorf microcentrifuge tube.Next, 600 µl of DTAB buffer (2% DTAB; 1.42 M NaCl; 20 mM EDTA; 100 mM Tris-ClpH 8.0; 2% PVP; 5 mM ascorbic acid; 4.0 mM Diethyldithiocarbamic acid ( DIECA)) was added to the sample and heated to 70°C.The prepared tubes were stirred using an Eppendorf Thermomixer comfort for 1 minute.The resulting mixture was added 500 µl of chloroform and isoamyl alcohol as a 24:1 mixture.All of this was stirred at 500 rpm for 5 minutes on an orbital shaker (ML-1).Next, centrifugation was performed for 5 minutes on an Eppendorf Centrifuge 5418 at 12,000 rpm, after which 200 µl of the upper liquid phase containing DNA was taken and placed in a sterile tube with 140 µl of isopropanol, stirred and left for 5 minutes at 18-20°C.After that, the sample was precipitated in a microcentrifuge at 12,000 rpm for 20 min, and then the supernatant was decanted and 800.0 µl of 70% ethanol was added.This was followed by resuspension and centrifugation for 5 minutes at 12,000 rpm.Subsequently, the supernatant was decanted again, precipitated for 2 minutes, and the ethanol residue was removed using a pipette.The precipitate was dried at 55°C and then DNA was dissolved in 100 µL TE at pH 8.0 (10 mM Tris-ClpH 8.0, 1 mM EDTA).After that, an electrophoregram was generated (Fig. 1).Spectrophotometric measurement of total plant DNA was then performed.Sample dilutions and controls were prepared.The sample was prepared as follows: 95 µl of deionised Milli-Q water (Millipore) and 5 µl of total DNA preparation were poured into a separate tube and mixed thoroughly.The control was treated as follows: 95 µl of water was poured into the tube, followed by 5 µl of TE buffer (pH 8).The concentration of the samples was then measured using a BioPhotometer (Eppendorf) v.1.35.The total DNA samples were then normalised to 30 ng/µL to measure their DNA concentration on a spectrophotometer.The results obtained made it possible to generate the following tabular data (Table 2).

Source: compiled by the authors
Then sea buckthorn DNA was diluted in TE buffer, which was a solution of 4 µg of DNA sample + 8 µg of PCR buffer with the HrMS014F/HrMS014R primer system.Then, electrophoresis was performed in a 2% agarose gel (1×LB buffer + ethidium bromide), and the electrophoregram showed the following image (Fig. 2).The field data were processed using Excel 2007 and Statistica 5.5.

RESULTS
The research carried out at the first stage of the study allowed to identify the forms of Hippophae rhamnoides L., most of which belong to bushy forms (Adaptyvna, Osoblyva, Mitsna, Sonyachne syayvo, etc.), the rest -to tree-like forms (F 1-10-11, F 1-2-500, F 2-12-4, F 8-18-32), as well as to distinguish forms by growth vigour and height, depending on the selection zone.It is worth noting that the bush forms (Morkvyana, Apel'synova, Adaptyvna, Nosivchanka) and some tree-like forms (F 1-10-11, F 2-12-4, F 1-2-500) are characterised by increased annual growth rates of about 0.90-1.10 and 0.70-1.30m, which is an important feature for nursery production.Osoblyva, Lymonna, and Sonyachne syayvo forms have a lower growth force, which allows for a reduction in the cost of manual labour for plant care when growing them in a collection nursery.
Forms Adaptyvna, F 1-2-500, and F 1-10-11 are characterised by dry detachment, annual fruiting, and, like forms Osoblyva and Nosivchanka, by the optimal ratio of polyphenolic compounds, organic acids, and sugars in fruits, which indicates their suitability for the production of healthy food.The sea buckthorn samples were registered as a working collection by the National Centre of Plant Genetic Resources of Ukraine for priority areas in breeding and genetic research (certificate No. 00292 dated 23.10.2020).Based on the breeding results, the best forms for economic traits were formed and identified, described, and systematised, taking into account the origin, morphological traits, biological characteristics, and biochemical properties.
The results of phenological observations allowed to find out that, in general, in the northern part of the Forest-Steppe of Ukraine, the beginning of the vegetation of sea buckthorn forms selected in the fallow ecosystems falls on the third decade of March -first decade of April, in Polissia -on the first decade of April.During the budding phase, the swelling and blooming of vegetative and generative buds are observed, which lasts on average 18 days (for Polissia conditions -about 21 days).However, the weather conditions of April 2022 led to the passage of certain stages of organogenesis in a shorter time, in particular, it was noted that the period from the beginning of bud development to the beginning of flowering lasted only 14 days, 7-8 days shorter than in 2019, 2020 and 2021.The deficit of precipitation in 2020 and 2021 during fruit filling caused a decrease in yield and fruit weight compared to 2019.The most stable forms by years in terms of productivity were F 1-2-500, Lymonna, Apelsynova, and less stable -Morkviana, F 1-10-11, Osoblyva (Fig. 3).The forms Adaptyvna, Osoblyva, F 1-10-11 and Lymonna were stable in terms of fruit weight (Fig. 4).

Figure 4. Berry mass of varieties and forms of Hippophae rhamnoides L. by years Source: compiled by the authors
Genetic and epigenetic differences between the forms of Hippophae rhamnoides L. are inherited, which causes gene expression under the influence of environmental factors, resulting in the manifestation of economically valuable traits, the identification of which is the basis for further breeding work.Sea buckthorn plants are characterised by low environmental resistance, in particular, when introduced to unfavourable soil and climatic conditions (high air temperatures, moisture deficit, heavy soils by mechanical composition, etc.), which causes the plants to become relatively weak in development and are affected by pathogens, damaged by pests, and often fall out, in particular, male forms.It should be noted that underestimation of these facts leads to a decrease in the productivity of female plants.The PCR analysis was used to trace the difference between the forms against the background of varieties to establish the range of relatedness and to identify the genetic polymorphism of the sea buckthorn genome under study.In the process of applying 5 DNA markers for molecular genetic characterisation of the studied culture, HrMS025 and HrMS026 were the most polymorphic, the others were also manifested in the studied samples, but mostly monomorphic (Fig. 5).It was found that the use of ISSR marker A17898 for sea buckthorn samples allows for qualitative determination of polymorphism and development of genetic passports (Fig. 7, Table 3).

Table 3. Certification of sea buckthorn genotypes
No.

Table 3, Continued
to be characterised by a high content of antioxidants and an inherent flavour-forming complex, which makes them suitable for the production of healthy food products.It should be added that several forms also have a high dry matter content, the share of which among other biochemical parameters varies between 18-25% (in particular, for forms F 1-15-6 and F 1-5-3).Fruits of forms F 1-15-8c, F 1-15-1, F 1-15-5a and F 1-15-2 contain higher than average soluble solids (>13%).
Based on the aforementioned, the of phenological observations, evaluation of breeding material for morphological traits, fruit biochemistry, and PCR analysis allowed us to select sea buckthorn forms with a set of valuable economic traits.This allowed to add new forms to the existing source material (Dublianska Osin, Prydorozhna, Pahorbova, Yaruzhna, Soborna, Adaptyvna Improved, Osinnya Krasunya, Rapsodiya), which were registered at the National Centre for Plant Genetic Resources of Ukraine, and the best ones (Soborna, Adaptyvna Improved, Osinnya Krasunya) received a certificate of registration from the General Bank of Ukraine.Sea buckthorn breeding samples are characterised by the following elements, in particular, the breeding form Yaruzhna, which is a seedling from free pollination of the Adaptyvna variety, has a dry fruit detachment, no or a small number of thorns, high resistance to fruit flies (9 b), high winter and drought resistance (9 b) and low shoot formation capacity.The breeding form Dublianska Osin, whose pedigree is a seedling from free pollination of the Adaptyvna variety, is characterised by dry fruit detachment, annual fruiting, late maturity, high resistance to fruit fly (9 b), high winter and drought resistance (9 b), high vitamin C content in fruits (70 mg/100 g), and high transportability (9 b).
The breeding form Prydorozhnia, which is a seedling from free pollination of Morkvyana variety, is characterised by high growth vigour (over 1.5 m/vegetation), productivity (total number of fruits per inflorescence -10 pcs.), high fruit yield (11 t/ha), annual abundant fruiting, medium early maturity, high winter hardiness (8 b), drought resistance (8.5 b), high content of vitamin C (72 mg/100 g), oil (3.8%), and has a universal type of fruit use.The breeding form Pahorbova is also a seedling from free pollination of the variety Morkvyana, characterised by high growth vigour (over 1.5 m/vegetation), productivity (number of fruits per inflorescence -10 pcs.), fruit yield (11 t/ha), annual fruiting, medium early maturity, high winter and drought resistance (9 b.), high content of vitaminC (83 mg/100 g) and carotene (11.5 mg/100 g), and has a universal type of fruit used.
The breeding form Adaptyvna improved, the pedigree of which is a seedling from free pollination of the Adaptyvna variety, has the following elements of novelty: moderate or medium growth vigour (up to 0.7 m/ vegetation), high productivity (increased number of fruits per inflorescence -10 pcs.), dry fruit detachment, absence or small number of thorns, early age of fruiting (5 years), annual fruiting, late maturity, high resistance to fruit fly (9 b), high winter and drought resistance (9 b), high vitamin C content (70 mg/100 g), high transportability and fruit tasting evaluation (9 b).The breeding form Rapsodiya is a seedling from free pollination of the variety Morkvyana and has the following novelty elements: high productivity (increased number of fruits per inflorescence -≥10 pcs.), high level of self-fruitfulness (≥75%), dry fruit detachment, low thorniness, high fruit yield (≥15 t/ha), annual fruiting, high vitamin C content (120 mg/100 g), universal type of fruit use, high suitability of fruits for blended cider production.
The Soborna breeding form is a seedling from free pollination of the Nadiina variety and is characterised by high productivity (number of fruits per inflorescence -10 pcs.), dry fruit detachment, increased fruit yield (15.6 t/ha), annual fruiting, thornlessness, high winter and drought resistance (9 b.), high vitamin C content (70 mg/100 g) and has a universal type of fruit used.The breeding forms Osinnia krasunia is a seedling from free pollination of the Morkvyana variety and is characterised by increased fruit yield (11 t/ha), dry fruit detachment, high resistance to fruit endomycosis (9 b), resistance to sea buckthorn fly (9 b), annual fruiting, late maturity, high winter and drought resistance (9 b) and fruit transportability (9 b).
Most of the new genotypes of female sea buckthorn have a tree life form (Dublianska Osin, Osinnya Krasunya, Soborna, Rapsodiya, Pahorbova, Pryrodorozhna, Yaruzhna), and one -Adaptyvna Improved -has a bush life form.Some forms (Adaptyvna Improved, Osinnya Krasunya, Soborna, Rapsodiya, Prydorozhnia) have an average growth force that does not exceed 1 m during the growing season, while the rest -Dublianskaia Osin, Yaruzhna and Pahorobova -have a growth force of about 1.5 m.All new forms have a semi-vertical position of branches, moderate crown density, average thickness of the annual shoot and the location of inflorescences on two-year-old and older shoots.It was found that Adaptyvna Improved and Soborna plants have no thorns, while the rest -Rapsodiya, Prydorozhna and Yaruzhna -have an average number of thorns.The leaf blade for all samples is narrow to broadly elliptical in shape, green with a faint silver bloom.Sea buckthorn samples differ in fruit shape and skin colour.For sea buckthorn, Dublianska Osin, Soborna, and Pryrozhna, the fruit shape is elongated-oval, Osnova Krasunya is broadly oval, Adaptyvna Improved is barrel-shaped, Rapsodiya, Pohorobova is elongated-barrel-shaped and for Yaruzhna it is rounded-barrel-shaped.The largest range of dark yellow, yellow-orange and dark orange shades is observed in Rapsodiya, Osinnya Krasunya, Pahorbova, and Pryrodozhna, while the rest ( Dublianska Osin, Adaptyvna improved, Soborna, Yaruzhna) have light yellow colour (Fig. 9, Table 4).In particular, to describe female forms of sea buckthorn, the system of ranking morphological parameters by the following features was improved (Table 4).

Table 4. Improved system for ranking morphological parameters of sea buckthorn by economically valuable traits
Fruit stalk by length, mm short (<3) average (from 3 to 5)  Note: for all forms, the flowering time is late; the content of polyphenolic compounds in fruits is average (301-500 mg/100 g of fresh weight); the content of titratable organic acids in fruits is average (4-5 mg/weight); the content of dry soluble matter in fruits is average (10-15 mg/weight), only for Adaptyvna Improved is it high (over 15 mg/weight); fruit transportability -high (8.0-9 points); fruit tasting score -average (6.5-8.0 points); frost, winter and drought resistancehigh, resistance to Fusarium wilt, fruit endomycosis, sea buckthorn fly -high (7.0-9 points); fruiting cycle -annual.Code for sea buckthorn forms: Dublianska Osin -1, Adaptyvna Improved -2, Osinnya Krasunya -3, Soborna -4, Rapsodiya -5, Pahorbova -6, Prydorozhnia -7, Yaruzhna -8, Lymonna -9 Source: compiled by the authors Almost all of the new varieties have a medium-long stalk, which makes them suitable for easier harvesting.Adaptyvna Improved, Yaruzhna and Dublianska Osin have a low fruit weight (up to 0.5 g), while Soborna has a high fruit weight.It is worth noting that most of the samples are characterised by an average content of ascorbic acid in the fruit, only the Rapsodiya sample has a high vitamin C content (over 100 mg%).The fruits of the Adaptyvna improved form have a high content of dry soluble matter compared to the others, for which this indicator is average.Adaptyvna improved, Pahorbova, Yaruzhna and Osinnya Krasunya have medium to high carotene content.The taste of the fruit in all forms is moderately sour or moderately sour and pleasant.All samples form medium and high yields and are characterised by high winter and drought resistance (9 points), high resistance to sea buckthorn fly, Fusarium wilt, and fruit endomitosis.Genotypes differ in ripeness, in particular, the time of the beginning of fruit ripening for Pridorozhna is early, Soborna, Rapsodiya and Pahorobova are medium, Adaptyvna Improved, Osinnya Krasunya and Yaruzhna are late, and Dublianska osin is very late, which is important for both breeding and production, in particular for staggered harvesting.
The new sea buckthorn varieties have proven to be productive and adaptive, in particular, the Adaptyvna Improved (UN3700088), Osinnya Krasunya (UN3700093), Soborna (UN3700090) and Rapsodiya (UN3700091) forms have been included in the Plant Genetic Resources of Ukraine and are promising source material, of which Adaptyvna Improved and Soborna will be submitted to the State Variety Testing in 2021.

DISCUSSION
The research and development of new varieties of sea buckthorn with high adaptability to changing climatic conditions, high productivity and exquisite fruit quality is an extremely important task to ensure stable yields, increase productivity and meet the needs of the population in food resources.Sea buckthorn culture is popular not only in Europe and Asia but also on other continents (Hakeem et al., 2018), due to the medicinal properties of the fruit.Therefore, the relevance of sea buckthorn culture obliges producers to grow varieties that meet modern requirements -characterised by high environmental stability, productivity, and fruit quality.In turn, such issues require the breeder to work hard and responsibly to develop appropriate source material for further breeding.
Sea buckthorn is believed to be a multipurpose medical crop (Gâtlan & Gutt, 2021).In particular, the anti-cancer activity of sea buckthorn raw materials has been proven.Much attention was also devoted to the antioxidant properties of polyphenols, flavonoids, and polysaccharides of sea buckthorn leaf and fruit extracts (Ma et al., 2019;Zhuo et al., 2019), including their processed products (juices, extracts, oils) (Ficzek et al., 2019).Today, many Ukrainians enjoy products made from sea buckthorn fruit and consider them a great food since the products of sea buckthorn processing do not lose their beneficial properties (Olas, 2018;Tkacz et al., 2020).Therefore, practitioners focus on the quality of the fruit, in particular the content of total sugars, organic acids (including malic acid), tannins, amino acids, trace elements, a complex of water-and fat-soluble vitamins, in particular ascorbic acid, and oil, as they are not sufficiently studied (Janceva et al., 2022).
Breeding for quality, in particular for high oil content, is important (Olas et al., 2018), as sea buckthorn oil has a positive effect on cell regeneration (Barkhuu et al., 2021), has a positive effect on oral health (Smida et al., 2019), is characterised by high antioxidant properties, is a source of bioavailable xanthophylls (Tudor et al, 2019), reduces blood cholesterol and changes the intestinal microbiota (Hao et al., 2019), and is noted for its cardioprotective, anti-atherogenic, antibacterial, antiviral and anti-inflammatory properties (Ulanowska et al., 2018).
According to experts (Leonov, 2022), sea buckthorn is just starting to be included in the ratings of crops popular with Ukrainian gardeners, but the global Table 4, Continued market for berries is growing every year (Rypan, 2023).Several Ukrainian farmers have assessed the prospects for sea buckthorn cultivation.At Rovy Agro (Kyiv region), sea buckthorn plants do not require large investments in protection and nutrition (AgroTimes, 2022).To maximise the biopotential of a sea buckthorn variety, proper environmental conditions are required to determine the productivity, composition, and quality of the fruit (Pintea & Magdas, 2022).
In breeding, methods of vegetative propagation, molecular breeding genetic transformants and invitro culture are used for mass reproduction of genotypes with economically valuable traits and preservation of genetic purity of hybrids (Bansal et al., 2018).Developments in sea buckthorn genomic resources have led to the creation of a variety of molecular markers, and DNA barcodes, which are important for targeted sea buckthorn breeding (Liu et al., 2022).In particular, the data today shows an increase in the resources of microsatellite markers.
Summarising the aforesaid, it can be stated that scientists are concerned with the impoverishment of sea buckthorn genetic diversity, therefore, in many countries, measures are underway to create genetic banks to preserve different genotypes belonging to subspecies subsp.caucasica, subsp.rhamnoides, carpatica, subsp.subsp.sinensis, subsp.turkestanica, and subsp.yunnanensis., subsp.mongolica, subsp.fluviatilis, subsp.wolongensis well adapted to specific conditions to involve them in the breeding process and obtain forms resistant to adverse environmental factors with high yield and fruit quality.
The results of a detailed study of sea buckthorn samples by morphological, physiological, and biochemical parameters of the fruit allowed to improve the methodology for systematising the methods of their description both during the route and stationary study, taking into account the origin, morphological characteristics, biological features, biochemical properties of the fruit.It has been shown that the best samples of Adaptyvna improved, Osinnya Krasunya, Soborna, Rapsodiya, Pahorbova, Prydorozhnia, etc. were transferred to the National Centre of Plant Genetic Resources of Ukraine for study, and due to high yield and fruit quality, Adaptyvna, Morkviana, Osoblyva and Nadiyna were transferred to the State Variety Testing, which resulted in certificates of registration of plant samples of the gene pool of Ukraine, plant genetic collection and certificates of State Registration of plant varieties suitable for distribution in Ukraine.For the first time in Ukraine, new sea buckthorn samples were genotyped in comparison with the best varieties, and the corresponding genetic passports were developed, which indicate genetic identity, which is important for breeding.
The second stage of research involved the involvement of sea buckthorn varieties Adaptyvna, Morkviana, Osoblyva, Nadiyna, etc. in breeding and the formation of 1.5-2 thousand hybrid nursery plants under controlled and uncontrolled pollination.The breeding work allowed us to study and select forms that are valuable in terms of a set of economic traits, such as Dublianska osin, Prydorozhna, Pahorbova, Yaruzhna, Osinnya Krasunya, Rapsodiya, etc.All samples were studied at the National Centre of Plant Genetic Resources of Ukraine, and the last four samples received a certificate of registration from the Ukrainian Plant Genebank.In 2021, the varieties Soborna and Adaptyvna Improved were submitted to the State Variety Testing.New high-yielding varieties of female sea buckthorn Adaptyvna, Osoblyva, Morkviana, Nadiina, Soborna, Adaptyvna Improved and pollinator varieties Obrii, Aboryhen adapted to the conditions of Polissia and Forest-Steppe of Ukraine have been proposed for production.
to the staff of the Institute of Horticulture of the National Academy of Agrarian Sciences of Ukraine for assistance in conducting agrotechnical activities in sea buckthorn nurseries, conducting biochemical analyses to determine the quality of the studied crop and processing of fruits to determine their suitability for the production of healthy food, as well as to the candidates of biological sciences Viktor Ryabchun and Olga Zadorozhna, head and leading presearcher of the National Centre of Plant Genetic Resources of Ukraine of the Plant Production Institute named after V.Ya.Yuriev of National Agrarian Academy of Sciences of Ukraine, for their work on thorough examination of applications and descriptions for sea buckthorn accessions, selection of the most valuable ones in the breeding and economic sense and their registration as accessions of the Genetic Bank of Ukraine.

Table 2 .
Results of measuring DNA concentration on a spectrophotometer