Nowadays, there is growing interest in new compositions and drugs that can preserve and maintain human health. There are many studies on both known probiotics and new sources of biologically active substances which may potentially be new avenues for human longevity. Here, we describe the protective effect of cell lysates of the relict bacteria Bacillus sp. F (Bacilus Lyuba), isolated from the ancient permafrost of Central Yakutia, upon DNA damage, induced by hydrogen peroxide. The DNA damage was monitored by a comet assay in leukocytes isolated from the peripheral blood of experimental animals (BALB/c mice). Cell lysates were prepared using the French Press and ultrasonic treatment. The levels of DNA damage in the presence of bacterial lysates upon the stress test (20 μM hydrogen peroxide for 10 min at 37°C) were almost twice lower, compared with the buffer control. Of note, the maximal protective effect was observed in the temperature range of 50°C–60°C; with the increase of the temperature, this effect vanished. The obtained results do not give a clear answer on the mechanism of the protective properties of Bacillus sp. F. (Bacilus Lyuba) lysates. To answer this question, a detailed analysis of the protein composition of bacterial lysate at different temperatures should be performed.

The effect of a coating material containing organic photoluminophore (PL) on the growth and development of mustard Brassica juncea L. plants colonized with beneficial associative bacteria Pseudomonas putida KT2442 and Rhodococcus erythropolis X5 was studied in vitro and in vivo. Plants grown with the use of microbial bacterization in combination with a photoluminophore coating (PLC) had significantly faster growth rates in vitro (2.1 times faster, P. putida; 1.8 times faster, R. erythropolis) than those grown using PLC alone (1.2 times faster). The leaves of plants grown with PLC had higher contents of glucose and fructose (28.4 ± 0.3% more glucose and 60.4 ± 0.3% more fructose accumulated compared to plants grown without PLC). It was found that seed weights and seed number increased 1.9-fold and 1.6-fold, respectively, for plants grown with PLC and colonized with beneficial P. putida KT2442 bacteria. The stimulatory effect of PLC on photosynthetic parameters of Photosystem II (PSII) was observed in colonized plants grown in vitro. For the first time, it was shown that providing plants with a PLC for only 4 weeks may make it possible to support further plant growth without PLC to obtain higher yields in the future. Thus, PLCs that convert shorter-wavelength radiation into red light may induce enhancement of biochemical processes not only in plants but also in microorganisms that supply plants with growth regulators and other active compounds. The results indicate the need for further research to understand the mechanisms of photobiological and photoregulatory systems in the interaction of microbes and plants.

Horticultural plants, in particular fruit trees, berry crops, and ornamentals, are valuable objects for studying their genetic and biochemical properties. Along with traditional methods of studying these plants, modern molecular genetic technologies are emerging, in particular genome editing using CRISPR/Cas9 nucleases. In this review, we have analyzed modern advances in genome editing of horticultural plants. To date, it has become possible to improve many plant characteristics using this technology, e.g., making plants resistant to biotic and abiotic stress factors, changing the time of flowering and fruit ripening, changing the growth characteristics of plants, as well as the taste properties of their fruits. CRISPR/Cas9 genome editing has been successfully carried out for many horticultural plants. Dozens of genes from these plants have been modified by means of genome editing technology. We have considered the main ways of delivering genetic constructs to plants as well as limitations that complicate the editing of target genes. The article reviews the prospects of using genome editing to improve the valuable properties of plants important to humans.

'Isabel' grape (Vitis labrusca x V. vinifera L. hybrid) is one of the main grape cultivars in Russia and some other countries for processing, due to its vigor, tolerance to the main fungal diseases, high yield and potential for sugar accumulation. The stilbene synthase gene VlvSTS was isolated from the hybrid grape cv. Isabel and cloned into a pSS plant transformation vector under the control of a constitutive 35S RNA double promoter of the cauliflower mosaic virus, CaMV 35SS. VlvSTS-gene containing transgenic tobacco lines were obtained and analyzed. For the first time plants expressing the VlvSTS gene were shown to have an enhanced resistance to the bacterial pathogen Erwinia carotovora subsp. carotovora B15. Transgenic plants were tested for resistance to a number of fungal pathogens. The plants were resistant to the grey mould fungus Botrytis cinerea, but not to the fungi Fusarium oxysporum, F. sporotrichioides, or F. culmorum. According to the results of a high performance liquid chromatography-mass spectrometry analysis, the amount of trans-resveratrol in leaves of transgenic plants with the highest expression of the VlvSTS gene was in a range from 150 to 170 μg/g of raw biomass. Change in the color and a decreased anthocyanin content in the flower corollas of transgenic plants were observed in transgenic lines with the highest expression of VlvSTS. A decrease in total flavonoid content was found in the flower petals but not the leaves of these tobacco lines. High expression of the VlvSTS gene influenced pollen development and seed productivity in transgenic plants. The size of pollen grains increased, while their total number per anther decreased. A decrease in the number of fertile pollen grains resulted in a decreased average weight of a seed boll in transgenic plants.

This work was aimed to study the opportunity of transgenic oilseed rape (Brassica napus L.), expressing the gene of antimicrobial peptide cecropin P1 (cecP1) to be inhabited with associative microorganisms Methylobacterium mesophilicum and Pseudomonas aureofaciens. Previously, resistance was demonstrated to the microbial pathogens:Erwinia carotovora B15, Pseudomonas syringae, Fusarium oxysporum, and Botrytis cinerea. Analysis of the fatty acid composition of transgenic plant seeds showed an increase in the proportion of unsaturated fatty acids. Plants studied showed increased photosynthetic activity in oxidative stress induced by paraquat instead of in oxidative stress and UV radiation compared to control ones. A study of the interaction of transgenic plants with the associative microorganisms showed that bacteria were located in all initially colonized plants, as well as in plants obtained after several passages of microproliferation. Accordingly, transgenic plants, containing the cecP1 gene were actively colonized by associative bacteria, indicating their safety for associative bacteria, indicating their safety for beneficial microorganisms.

Ectopic auxin overproduction in transgenic potato leads to enhanced productivity accompanied with concerted and occasional changes in hormonal status, and causing altered response of transformants to exogenous auxin or cytokinin.

Kalanchoe pinnata L. plants bearing an artificial CP1 gene encoding the cecropin P1 antimicrobial peptide have been obtained. The presence of the CP1 gene in the plant genome has been confirmed by PCR. Cecropin P1 synthesis in transgenic plants has been shown by MALDI mass spectrometry and Western blotting. The obtained plants have been highly resistant to bacterial and fungal phytopathogens, and their extracts have demonstrated antimicrobial activity towards human and animal pathogens. It has been shown that transgenic plants bearing the CP1 gene can be colonized by the beneficial associative microorganisms Methylovorus mays.

Kalanchoe pinnate (Kalanchöe pinnata L. ) plants with synthetic gene of antimicrobial peptide cecropin P1 (CP1) under the control of promoter 35S RNA of cauliflower mosaic virus (CaMV 35S) were produced. For transformation, a modified binary vector not containing selective genes of tolerance against antibiotics and herbicides was used. Screening of the marker-free transformed plants was conducted on the medium without selective antibiotics by revealing antibacterial activity of plant extracts and cecropin P1. The marker-free plants produced displayed increased resistance against bacterial and fungus phytopathogens, while their extracts were characterized by antimicrobial activity for human and animal pathogens. These plants meet the requirements of biosafety and may be used as producers of cecropin P1 in pharmaceutics.

Ген антимикробного пептида цекропина Р1 (СP1) встраивали в векторную плазмиду pPCV91 под контролем промотора 35S РНК вируса мозаики цветной капусты (CaMV 35S), содержащего 4 энхансерных последовательности CaMV 35S и нетранслируемую лидерную последовательность РНК вируса табачной мозаики. С помощью полученного рекомбинантного вектора проведена агробактериальная трансформация растений табака (Nicotiana tabacum L.) сорта Самсун. Присутствие гена СР1 в геноме растений подтверждено методом полимеразной цепной реакции. Экспрессия гена СР1 в трансгенных растениях доказана вестерн-блот анализом и тестированием антибиотической активности растительных экстрактов. Уровень синтеза цекропина Р1 в различных линиях составлял 0.02–0.2% от общего растворимого белка листьев растений. Трансгенные растения проявляли по сравнению с контрольными растениями повышенную устойчивость к фитопатогенным микроорганизмам и к окислительному стрессу. Показано, что способность трансгенных растений экспрессировать цекропин Р1 передавалась потомству.

A gene of antimicrobial peptide cecropin P1 (CP1) was inserted into the vector plasmid pPCV91 under the control of promoter 35S RNA of cauliflower mosaic virus (CaMV 35S) containing four enhancer sequences CaMV 35S and nontranslated leader sequence Ω RNA of tobacco mosaic virus. The recombinant vector obtained was used for agrobacterial transformation of tobacco plants (Nicotiana tabacum L., variety Samsun) with the polymerase chain reaction (PCR)-based method. The presence of gene CP1 in the genome of plants was proven by western-blot analysis and testing the antibiotic activity of plant extracts. In different plant lines, the level of cecropin P1 synthesis amounted 0.02–0.2% of total soluble plant leaf protein. The transgenic plants, unlike the control ones, displayed enhanced tolerance to phytopathogenic microorganisms and oxidative stress. It was established that the ability of the transgenic plants to express cecropin P1 is transmitted to progeny.

Oral immunogenicity of the hepatitis B surface antigen (HBsAg) synthesized in the tubers of marker-free potato plants has been demonstrated. Experiments were performed in the two groups of outbred NMRI mice. At the beginning of investigations, the mice of experimental group were fed the tubers of transgenic potato synthesizing the HBsAg three times. The mice of control group were fed nontransgenic potato. Intraperitoneal injection of the commercial vaccine against hepatitis B (0.5μg/mouse) was made on day 71 of the experiment. Enzyme-linked immunoassay (ELISA) of the serum of immunized animals showed an increase in the level of HBsAg antibodies significantly above the protective value, which was maintained for 1 year after the immunization. In 1 year, the experimental group of mice underwent additional oral immunization with HBsAg-containing potato tubers. As a result, the level of antibodies against the HBsAg increased and remained at a high protective level for several months. The findings show the possibility of using transgenic plants as a substance for obtaining a safe edible vaccine against hepatitis B.

Phytohormones, auxins in particular, play an important role in plant development and productivity. Earlier data showed positive impact of exogenous auxin on potato (Solanum tuberosum L.) tuberization. The aim of this study was to generate potato plants with increased auxin level predominantly in tubers. To this end, a pBinB33-tms1 vector was constructed harboring the Agrobacterium auxin biosynthesis gene tms1 fused to tuber-specific promoter of the class I patatin gene (B33-promoter) of potato. Among numerous independently generated B33:tms1 lines, those without visible differences from control were selected for detailed studies. In the majority of transgenic lines, tms1 gene transcription was detected, mostly in tubers rather than in shoots. Indoleacetic acid (IAA) content in tubers and the auxin tuber-to-shoot ratio were increased in tms1-expressing transformants. The organ-specific increase in auxin synthesis in B33:tms1-transformants accelerated and intensified the process of tuber formation, reduced the dose of carbohydrate supply required for in vitro tuberization, and decreased the photoperiodic dependence of tuber initiation. Overall, a positive correlation was observed between tms1 expression, IAA content in tubers, and stimulation of tuber formation. The revealed properties of B33:tms1 transformants imply an important role for auxin in potato tuberization and offer prospects to magnify potato productivity by a moderate organ-specific enhancement of auxin content.

The review considers the basic strategies used to produce biologically safe marker-free transgenic plants and analyzes their advantages and disadvantages. The systems of positive and negative selection as safer approaches for transformant identification are briefly described. The application of co-transformation, transposition, and site-specific recombination for production of marker-free plants is described. Special attention is paid to novel approaches to create marker-free plants initially containing no selective genes in their genomes.

The arbuscular mycorrhiza development has been studied in transgenic tobacco plants expressing an agrobacterium iaa M gene of auxin biosynthesis (iaaM-plants) and plants with a heterologous gene of isoprenoid biosynthesis pathway hmg1 from Arabidopsis thaliana in the sense and antisense orientations relative to the promoter (hmg1and as-hmg1-plants, respectively). The mycorrhiza occurrence frequency has been shown to be higher in iaaMand hmg1-plants as compared to control and as-hmg1-plants.

В обзоре рассмотрены основные стратегии получения биологически безопасных безмаркерных трансгенных растений и проанализированы их достоинства и недостатки. Кратко описаны системы позитивной и негативной селекции как более безопасные подходы выявления трансформантов. Описано использование методов ко-трансформации, транспозиции и сайт-специфической рекомбинации для получения безмаркерных растений. Особое внимание уделено новым подходам к созданию безмаркерных растений, изначально не содержащих селективных генов в своем геноме.