All Russia Research Institute of Agricultural Biotechnology

Moscow, Russia

All Russia Research Institute of Agricultural Biotechnology

Moscow, Russia
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Miroshnichenko D.,RAS Institute of Basic Biological Problems | Chaban I.,All Russia Research Institute of Agricultural Biotechnology | Chernobrovkina M.,All Russia Research Institute of Agricultural Biotechnology | Dolgov S.,RAS Institute of Chemistry
PLoS ONE | Year: 2017

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. © 2017 Miroshnichenko et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Lazareva E.M.,Moscow State University | Baranova E.N.,All Russia Research Institute of Agricultural Biotechnology | Smirnova E.A.,Moscow State University
Cell and Tissue Biology | Year: 2017

We examined the organization of microtubule system of interphase cells in roots of Medicago sativa L. during acclimation to salt and osmotic stress at different concentrations of NaCl, Na2SO4, and mannitol. We identified morphological changes of tubulin cytoskeleton in different root tissues during the acclimation to salt and osmotic stress: (1) decreased density of the cortical microtubule network, (2) random orientation of cortical microtubule bundles, (4) thickening of the bundles, (3) nonuniform density of the bundles, (4) fragmentation of the bundles, and (5) formation of microtubule converging centers. Network thinning and thickening of the bundles were observed both under osmotic and salt stress. Random orientation of cortical microtubules was visualized under osmotic stress but not during salt stress. Fragmentation of microtubule bundles took place under salt stress with a high concentration of mannitol. Formation of microtubule converging centers was common under prolonged action of sodium sulfate, less evident under sodium chloride, and not found after mannitol treatment. Our data show that, in alfalfa root cells, cortical microtubules rearrange not only in response to different ions, but also to osmotic pressure. Thus, the signaling pathways and molecular mechanisms inducing reorganization of the microtubule system may be triggered by sodium cations, as well as by sulfate and chloride anions at concentrations that do not cause irreversible cell damage. © 2017, Pleiades Publishing, Ltd.


Sochivko D.G.,JSC Syntol | Fedorov A.A.,Russian Academy of Sciences | Alekseev Y.I.,All Russia Research Institute of Agricultural Biotechnology | Kurochkin V.E.,Russian Academy of Sciences | Dubina M.V.,St Petersburg National Research Academic University
Doklady Biochemistry and Biophysics | Year: 2017

The course of the real-time polymerase chain reaction (PCR) is determined by the temperature dependence of the kinetics of the component reactions, particularly the DNA strand hybridization. To investigate the effect of thermal processes on the reaction behavior, a mathematical model in which the variable rate constant of dissociation of “primer–single strand” complexes depends on temperature was proposed. The reaction medium temperature, which depends on time, was also introduced into the model. The proposed model of real-time PCR makes it possible to analyze different aspects of the reaction, which are important for the development of instruments and reagents for PCR. © 2017, Pleiades Publishing, Ltd.


Zlobin N.E.,All Russia Research Institute of Agricultural Biotechnology | Ternovoy V.V.,All Russia Research Institute of Agricultural Biotechnology | Grebenkina N.A.,All Russia Research Institute of Agricultural Biotechnology | Taranov V.V.,All Russia Research Institute of Agricultural Biotechnology
Vavilovskii Zhurnal Genetiki i Selektsii | Year: 2017

There are several technologies for plant genome editing, of which the most simple and universal is CRISPR/ Cas. Currently, this technology is widely used for gene knockout, deleting genome fragments and inserting exogenous sequences in the plant genome. For each of these applications, many different types of genetic tools have been developed that are used by various research groups to solve specific problems. The CRISPR/Cas technology for plant genome editing is at an early stage of optimization, which is reflected by the ongoing search for the most effective, simple and flexible techniques. As a result, experimental work has to be preceded by a rather long and laborious process of selecting a genetic tool that will be optimal for a specific experimental task. In our review we describe the main variants of the CRISPR/Cas technology used to edit a plant genome. We classify them in terms of experimental tasks solved, major components and technology performance. In the first half of the review a detailed description of two major components of CRISPR/Cas technology - nuclease and guide RNA - is given, the effect of structural features of these elements on editing efficiency is analyzed. Experimental data on the relationship between editing efficiency and nucleotide sequence of guide RNA are generalized. We also give the characteristic for different variants of nucleases used for plant genome editing and discuss their benefits for different experimental purposes. In the second half of the review various strategies for expression of CRISPR/Cas elements in plant cells, in particular, advantages and disadvantages of stable transformation and transient expression, are discussed. The effect of various regulatory elements of genes encoding nuclease and guide RNA on editing efficiency is described. Special emphasis is placed on the techniques of increasing targeted gene replacement efficiency. © AUTHORS, 2017.


Rogozina E.V.,Federal Research Center The Ni Vavilov All Russian Institute Of Plant Genetic Resources Vir | Khavkin E.E.,All Russia Research Institute of Agricultural Biotechnology
Vavilovskii Zhurnal Genetiki i Selektsii | Year: 2017

Interspecific hybridization is the primary method of potato breeding for resistance to pathogens. By introgressing genetic material from various species of the genus Solanum L. and selecting the best combinations, it is possible to merge, in a single genotype, both high productivity and resistance to diseases and pests. Among the most impressing outcomes of potato breeding, we find varieties resistant to late blight, potato virus Y and nematodes. In the pedigrees of many recent varieties (breeding lines) that are resistant to pathogens of different nature (fungi, oomycetes, bacteria or viruses), we find genetic material of Solanum andigenum, S. demissum, S. stoloniferum, S. acaule, S. vernei and other wild and cultivated potato species. The breeding value of tuber-bearing Solanum species depends on their compatibility with the cultivated potato and the mechanism of target trait inheritance. To overcome incompatibility in crosses, breeders employ ploidy manipulation in the parental forms, bridge crosses, the mediator method and various in vitro technologies. Potato genotypic variation is significantly expanded by interspecific hybridization of wild and cultivated potato relatives. The main components of breeding technology based on interspecific potato hybrids are the identification of promising initial genotypes, the control over the introgression of the target traits through the crosses, the selection of hybrid clones prospective for practical breeding and the assessment of their donor properties. All of these processes are greatly accelerated and promoted by the methods of marker-assisted selection. Advanced lines and clones developed by interspecific hybridization manifest a high diversity of genotypes and particular genes and are unique donors for breeding new varieties maintaining high and durable resistance to diseases and pests. © AUTHORS, 2017.


Miroshnichenko D.,RAS Shemyakin Ovchinnikov Institute of Bioorganic Chemistry | Chernobrovkina M.,All Russia Research Institute of Agricultural Biotechnology | Dolgov S.,RAS Shemyakin Ovchinnikov Institute of Bioorganic Chemistry
Plant Cell, Tissue and Organ Culture | Year: 2016

The efficiency of immature embryo-derived in vitro culture of G genome wheats is significantly influenced by various auxins and sugars which are used for induction of embryogenic response, and by regeneration media composition for promotion of plant development from subcultured embryogenic calli. The embryogenic calli of Triticum timopheevii has demonstrated the highest regeneration ability when the initial explants were cultured on the media supplemented with 4 mg l−1 of Picloram (29.0 %), 4 mg l−1 of Dicamba (28.7 %) or 3 mg l−1 of 2,4-D (29.1 %). The media supplemented with 5–6 mg l−1 of Picloram were considered to be the most effective for promotion of embryogenic/regenerable callus production in Triticumkiharae cultures (73.7–75.0 %). Both T. timopheevii and T. kiharae embryogenic structures were characterized by the formation of green and albino plantlets. Generally the medium that was initially supplemented with Picloram promoted the formation of lower albino plants fraction rather than 2,4-D and Dicamba. As it was measured by the total green plant production per initial explant, the overall efficiency has been reduced when sucrose was substituted by glucose or maltose. The regeneration medium supplemented with 0.25 mg l−1 TDZ significantly enhanced the regeneration capacity of embryogenic callus in T. kiharae. In culture of T. timopheevii the difference between the medium lack of growth regulators and the medium supplemented with TDZ was not prominent, though both of the media have demonstrated the greater efficacy as compared to those supplemented with BA and Zeatin. © 2016 Springer Science+Business Media Dordrecht


PubMed | Russian Academy of Sciences, JSC Syntol, Federal Biomedical Agency and All Russia Research Institute of Agricultural Biotechnology
Type: | Journal: Doklady. Biochemistry and biophysics | Year: 2016

The paper reviews different approaches to the mathematical analysis of polymerase chain reaction (PCR) kinetic curves. The basic principles of PCR mathematical analysis are presented. Approximation of PCR kinetic curves and PCR efficiency curves by various functions is described. Several PCR models based on chemical kinetics equations are suggested. Decision criteria for an optimal function to describe PCR efficiency are proposed.


PubMed | Russian Academy of Sciences, JSC Syntol, Federal Biomedical Agency and All Russia Research Institute of Agricultural Biotechnology
Type: Journal Article | Journal: Doklady. Biochemistry and biophysics | Year: 2016

Macroscopic kinetic models describing the process of polymerase chain reaction (PCR) are currently solved only by numerical methods, which hampers the development of effective software algorithms for processing the results of the reaction. This paper considers the application of the homotopy perturbation method for obtaining approximate analytical solution of the simplest system of enzymatic kinetic equations describing the synthesis of nucleic acid molecules during PCR. The resulting approximate analytic solution with high accuracy reproduces the results of a numerical solution of the system in a wide range of ratios of enzyme and substrate concentrations both for the case of a large excess of the substrate over the enzyme and vice versa.


PubMed | Russian Academy of Sciences, JSC Syntol, Federal Biomedical Agency and All Russia Research Institute of Agricultural Biotechnology
Type: Journal Article | Journal: Doklady. Biochemistry and biophysics | Year: 2017

Development of methods for obtaining approximate analytical solutions of nonlinear differential equations and their systems is a rapidly developing field of mathematical physics. Earlier, an approximate solution of the simplest system of kinetic enzymatic equations for calculating dynamics of complementary strands of nucleic acids was obtained. In this study, we consider an alternative approach to selecting the basic linear approximation of the used method, which makes it possible to obtain more accurate analytical solutions of the set problem.


PubMed | All Russia Research Institute of Agricultural Biotechnology and IBC Generium LLC
Type: Journal Article | Journal: BMC biotechnology | Year: 2016

In a previous study we found that in chickweed the expression level of the pro-SmAMP2 gene was comparable or even higher to that of the -actin gene. This high level of the gene expression has attracted our attention as an opportunity for the identification of novel strong promoters of plant origin, which could find its application in plant biotechnology. Therefore, in the present study we focused on the nucleotide sequence identification and the functional characteristics of the pro-SmAMP2 promoter in transgenic plants.In chickweed (Stellaria media), a 2120bp promoter region of the pro-SmAMP2 gene encoding antifungal peptides was sequenced. Six 5-deletion variants -2120, -1504, -1149, -822, -455, and -290bp of pro-SmAMP2 gene promoter were fused with the coding region of the reporter gene gusA in the plant expression vector pCambia1381Z. Independent transgenic plants of tobacco Nicotiana tabacum were obtained with each genetic structure. GUS protein activity assay in extracts from transgenic plants showed that all deletion variants of the promoter, except -290bp, expressed the gusA gene. In most transgenic plants, the GUS activity level was comparable or higher than in plants with the viral promoter CaMV 35S. GUS activity remains high in progenies and its level correlates positively with the amount of gusA gene mRNA in T3 homozygous plants. The activity of the ro-SmAMP2 promoter was detected in all organs of the transgenic plants studied, during meiosis and in pollen as well.Our results show that the ro-SmAMP2 promoter can be used for target genes expression control in transgenic plants.

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