Components Of Nutrient Medium Research Articles

Research on in vitro propagation techniques of vine-tea plant (Ampelopsis cantoniensis (Hook. and Arn.) Planch)

Vine-tea plant has the scientific name Ampelopsis cantoniensis (Hook. and Arn.) Planch and is a valuable herb plant in Vietnam. This study aimed to find a method to sterilize samples and nutrient medium components in each stage of shoots regeneration, rapid shoots multiplication, complete plant creation and in vitro seedling cultivation in the nursery stage of in vitro propagation techniques of vine-tea plant. Four experiments in research room and one experiment in nursery were conducted from January to December 2023 at Hue University of Agriculture and Forestry, Vietnam. Criterias of infected, alived, clean and dead samples, rate of shoot regeneration and number of shoots/samples were measured. Research results showed that the best medium to regenerate shoots from stem section carrying dormant eyes is MS + 1.5mg BAP/l + 6.5g agar/l + 30g sucrose/l. The best medium to quickly multiply shoots is MS + 1.5 mg BAP/l + 0.2 mg IBA/l + 6.5g agar/l + 30g sucrose/l. The best complete plant-creating medium is MS + 0.5 mg α-NAA/l + 6.5g agar/l + 30g sucrose/l. In the condition that the nursery has a rain cover roof, a black net reduced the light intensity by 70% and the tissue culture nursery substrate of humus soil under the forest canopy + coconut fiber powder + smoked rice husk (1:1:1) gave the best results with survival rate reached 76.67%.

Влияние концентрации и рецептуры макросолей в питательной среде на показатели регенерации меристем, на этапе ввода в культуру in vitro, у некоторых аборигенных донских сортов винограда

Abstract. Rare indigenous grape varieties are of significant value for viticulture, breeding and preservation of genetic material. Often, ancient grape varieties, due to prolonged vegetative propagation, are infected with one or more chronic diseases (viruses, phytoplasmas, bacterial cancer). The most reliable way to improve the health of such plants is the method of apical meristems in in vitro culture. In which, on artificial nutrient medium, it is necessary to regenerate the plant from isolated extremely small apical explants (about 50-100 microns). One of the most important components of nutrient medium are macroelements. Not only their quantity is of great importance, but also the compounds they represent, as well as the balance and ratio of all elements in the nutrient medium. In this regard, the purpose of our study was to study the characteristics of mineral nutrition of grape plants of native Don varieties in in vitro culture. To do this, we studied the effectiveness of the most commonly used nutrient media in microclonal propagation. The experiments were carried out on four rare native Don varieties Plechistik oboepolyi, Bessergenevskiy №10, Musketnyi and Durman. Protocols used: Murashige and Skuug (MS), McCown Wood Plant (WPM), Chee and Pool (C2D), Quoirin & lepoivre (QL) and Anton Rebrov (AR1). The nutrient media differed only in the composition of macrosalts; all other components were added in equal quantities. The varietal response of the studied varieties was different. The Plechistik oboepolyi and Bessergenevskiy №10varieties regenerated best, and the Musketnyi and Durman varieties regenerated more poorly. It has been established that the composition of macroelements significantly affects the regeneration and differentiation of shoots in apical meristems. For most varieties, the most stable results were observed when using WPM and AR1 nutrient media. The worst development of explants in all varieties in the study occurred on MS and C2D media. Key words: apical meristem, introduction into in vitro culture, regeneration of explants, recipes for nutrient media, concentration of macronutrients, native Don varieties

Clonal micropropagation of apple trees

The purpose of the review is to analyze the state of knowledge of the issue of apple tree reproduction in Russia and abroad. Of all the methods of tissue culture, clonal micropropagation is the most studied, which has a number of advantages in comparison with traditional vegetative multiplication: obtaining healthy material, rapid reproduction, obtaining genetically homogeneous material, the possibility of releasing material throughout the year. Micropropagation has played an important role in the production of healthy, disease-free plants and in the rapid multiplication of apple varieties and rootstocks. This review shows that over the past decades, foreign and domestic researchers, including scientists of our institute, have developed a large number of methods for cultivating of varieties and rootstocks of apple trees. Due to the heterozygosity of the genus Malus, protocols for specific gene types are often developed. The conducted research analysis showed that the successful reproduction of apple trees is influenced by external and internal factors (genotype, physiological state, components of nutrient media, conditions of adaptation of micro-plants). The review reflects various approaches to solving such problems as obtaining an aseptic culture at the initiation stage, reducing the negative effect of phenol oxidation products, increasing the proliferative activity of explants using various nutrient media, as well as natural and synthetic growth hormones - cytokinins, problems of stimulating root formation as a result of the use of various types of auxins and methods of exposure, the development of conditions for the adaptation of microplants of varieties and rootstocks of apple trees in vivo. The emergence of new varieties will require the development of new cultivation conditions.

Laccase produced by Coriolopsis trogii and Cerrena unicolor with the mixed of metal ions and lignocellulosic materials

Coriolopsis trogii and Cerrena unicolor were investigated for laccase production in submerged fermentation with different nutrient medium containing metal ions and lignocellulosic materials. The maximum laccase activity of C. trogii Han751 was 8584.44 ± 98.45 U/L and was obtained from nutrient medium 7. However, the maximum laccase activity of C. unicolor Han 849 was 16144.26 ± 635.30 U/L from nutrient medium 9. Thus, the capacity of secreting laccase of C. unicolor Han 849 was superior to that of C. trogii Han751. Different fungal species have different medium components suitable for laccase production. The content of CuSO4·5H2O and MnSO4 in nutrient medium with the concentration of 0.25 g/L and 0.151 g/L, respectively, was more beneficial to C. trogii Han751 secreting laccase. However, the vital components of nutrient medium that contribute to the laccase activity of C. unicolor Han 849 were corncob, glucose, and CuSO4·5H2O, with the corresponding concentrations of 1 g/flask, 5 g/L, and 0.25 g/L, respectively. The results will contribute to the development of new methods to produce low-cost laccase.

Electrophysical sensor systems for in vitro monitoring of bacterial metabolic activity

Intense use of microbial cells in biotechnological production requires rapid methods to monitor bacterial growth online and analyze the regulation of microbial metabolic activity. Some details of metabolic activity are examined conventionally, by changing the concentration of nutrient medium components, the concentration of metabolites, the pH, the respiratory activity, and other variables. In this case, the set of variables used may turn out insufficient and a number of critical points of the control process will remain in the shadow. The enzymatic catalysis in the bacterial cell is accompanied by a redistribution of charges inside the cell and, therefore, may lead to a change in its electrophysical characteristics. Ideally, researchers should use structurally intact and metabolically undisturbed growing cells to fully understand the enzymatic processes in microbes. Electrophysical analysis, based on the study of cells as electrophysical objects and on the measurement of their polarization characteristics, is a new approach to assessing the intravital physiological variables of cells and their heterogeneity. This article describes the theoretical principles of electrophysical analysis of bacteria and shows the possibilities of methods for examining the metabolic status of bacteria.

Improvement of clonal micropropagation technique of promising Lonicera caerulea L. cultivars

The work is focused on enhancement of in vitro propagation technique of valuable Lonicera caerulea L. cultivars. The impact of some nutrient medium components at main cultivation stages of the genus Lonicera L. representatives was studied. Some cultivars showed the positive effect of substituting sucrose for glucose in the medium at the multiplication stage (Diana (30 or 40 g L−1), Moskovskaya 23 (20 g L−1) and Yugana (40 g L−1)). When studying the influence of different auxin types on honeysuckle rhizogenesis, Solovey and Yugana preferred addition of 1.0 mg L−1 IBA into the medium. During the research of influence of different auxin concentrations on L. caerulea rooting the increased number of rooted microshoots of Goluboy Desert was demonstrated with the increasing of IAA concentration (from 0.5 to 3.0 mg L−1). Meanwhile, Diana and Yugana better rooted on the medium containing 1.0 mg L−1 IAA but Zolushka reached its highest rooting percentage on the medium supplemented with 0.5 mg L−1 IAA. The assessment of second-year plants undergone the whole cycle of clonal micropropagation was carried out 85% of these plants began to bear fruits.

OPTIMIZATION OF NUTRIENT MEDIUM COMPOSITION FOR CULTIVATION OF CONSORTIUM OF STRAINS BACILLUS VELEZENSIS BIM В-497 D AND BACILLUS VELEZENSIS BIM В-1312 D

Stepwise optimization of nutrient medium components was carried out for bacterial consortium B. velezensis BIM В-497 D and B. velezensis BIM В-1312 D as the basic of feed additive Bacicorn. The appropriate carbon source – corn meal was selected and its optimal concentration was determined. Basic composition experiment was conducted with subsequent sliding along concentration gradient. Cell titer CFU/ml was used as optimization parameter. Optimization increased 2,8 times the
 level of viable cells of bacterial consortium in the cultural liquid.

Significance of nutrient media choice for the long-term cultures of leukemic T-lymphoblasts

Correct choice of nutrient media for culturing different types of cells in various applications is one of the most important aspects of modern biotechnology, since chemical composition of the culture media largely contains the necessary metabolites to support certain cells’ growth lines outside the body. Jurkat line of human leukemic T-lymphoblast-like cells (hereinafter Jurkat T-cells) is actively used for in vitro modeling of intracellular signaling and activation of normal blood T-lymphocytes mediated by the T-cell receptor/CD3/ CD4 complex in toxicological studies of immune and secretory responses, to test medicinal substances and ions. Also, Jurkat T-cells are widely used for ex vivo testing in immunology, oncology, toxicology, orthopedics, and traumatology. The existing standards and numerous studies are mainly based on short-term in vitro cultivation of Jurkat T-cells in RPMI 1640 nutrient medium. Meanwhile, the issues of long-term maintenance of the growth of Jurkat T-cells culture are poorly presented in the research literature. This study aimed for studying the activity of Jurkat T-cells over 7 to 14 days of in vitro culture and comparing the relative value of RPMI 1640 and αMEM media for the behavior of immunocompetent tumor cells. Using flow cytometry, multiplex analysis, and phase contrast Cell-IQ microscopy, the proportions of living cells and those dying by apoptosis and necrosis, secretion of cytokines and chemokines, and the dynamics of cell biomass propagation were studied. It was found that the αMEM medium in the complete nutrient medium, as compared with RPMI 1640, is more appropriate to in vitro promotion of cell viability (increased proportion of viable cells by 13.5% at the day 14), their secretory ability for 23 из 27 tested biomolecules, shortened adaptation time (на 32%) in culture before growth initiation, 5-fold increase of the Jurkat Т-cell cellularity by the day 7. Potential significance of the chemical components of nutrient media and secreted biomolecules for these results is discussed. As based on the results obtained, we concluded on superior properties of αMEM medium for long-term in vitro cultures of Jurkat T-cells. Consequently, the in vitro testing of medical devices intended for long-term contact with the body, including those for cancer patients, using Jurkat T-cell leukemia line in RPMI 1640 medium, may lead to wrong predictions on their biocompatibility and potential antitumor activity.

Increase of bacteriocin-like activity of <i>Bacillus thuringiensis</i> strain by improving nutriculture medium composition

The development of the nutriculture medium composition and the optimal concentrations of its constituent components to increase the bacteriocin-like activity of the strain Bacillus thuringiensis ssp. Dakota are presented. The study was carried out using a multifactor experiment with further processing of statistical data in order to optimize the basic nutriculture medium and maximize the activity of the target product. Sources of nitrogen (peptone and yeast extract) and carbon (glycerin and glucose) were used as optimization factors. The control growth medium was medium "A", traditionally used for the cultivation of bacteria of the genus Bacillus. The degree of impact of the studied factors on the effective feature was determined using the model of multiple linear regression of the first order, whereby the optimal ratio of the components was calculated on the basis of the quadratic model. The ability of the Bacillus thuringiensis ssp. dakota strain to produce a bacteriocin-like substance (BLIS) was revealed. The dependence of BLIS synthesis on the cultivation medium was established: on the carbohydrate-free medium, the antimicrobial activity of BLIS was 1.5 times lower than that on the medium containing glucose and glycerin. To obtain maximum BLIS activity, the carbohydrate medium was optimized using a multifactor experiment performed by the method of orthogonal Latin rectangles. Mathematical models of linear regression of the first and second order were constructed depending on the concentration of nutrient medium components. The optimal concentration of the components was determined on the basis of a second-order regression model that takes into account the effects of the interaction of factors and the nonlinearity of the process. As a result of optimization of the nutrient medium, the quantitative composition of the components of the culture medium was determined: peptone – 9 g/l; yeast extract – 2.6; glycerol – 5.6 g/l, on which the antimicrobial activity of BLIS increased by 60% compared with the activity on the initial medium. It was shown that the synthesis of BLIS, unlike the synthesis of delta-endotoxin, is not regulated by catabolite repression of carbon.

THE FEATURES OF THE ACCUMULATION OF FLAVONOIDS IN BIOTECHNOLOGICAL RAW MATERIAL OF IRIS SIBIRICA L. THE DEVELOPMENT OF METHODS OF QUANTIFICATION

Identification of scientific regularities of accumulation of physiologically active compounds is relevant to the issues of plant biology, as it can create conditions for the rapid development of biotechnological approaches and the solution of a number of environmental and economic problems associated with the use of plant raw materials. The aim of this work was to identify the features of the accumulation of flavonoids and the development of methods of differential spectrophotometry allows to assess the quality of biotechnological raw materials Iris sibirica L. (Siberian iris) on the content of flavonoids.
 The most important regulators of the synthesis of secondary compounds in plant tissue cultures include such components of nutrient media as hormones. As a result of our experiments it was noted for I. sibirica characteristic relationship between the accumulation of biomass and the content of quercetin and rutin. On the medium with 5.0 µm BAP, supplemented by auxins with an increase in the total height of the shoots, the quercetin content and rutin in the phytomass decreased dramaticly. To maintain a balance between biomass accumulation and flavonoid content for I. sibirica, we recommend using media with 2.5 µm BAP supplemented with auxins.
 The developed method allows to determine the content of the total of flavonoids in Iris sibirica raw materials in the presence of other compounds, is simple in execution and does not require expensive equipment. A validation evaluation of the technique indicates its suitability for quality control of biotechnological raw materials Iris sibirica.

Optimization of nutrient medium components for production of a client endo-β-1,4-xylanase from Aspergillus fumigatus var. niveus using a recombinant Aspergillus nidulans strain

The goal of this study was to optimize a nutrient medium to maximize production of endo-β-1,4-xylanase (hereafter referred to as xylanase) using an Aspergillus nidulans modified by integration of an AFUMN-GH10 gene from Aspergillus fumigatus var. niveus. This modification resulted in high-yield secretion and accumulation of recombinant protein. Xylanases are an industrially relevant enzyme class used in food production and bioprocessing. High-titer production of xylanase was achieved by the cultivation of A. nidulans in the presence of maltose. A 2-level Plackett-Burman design was used to determine which medium components significantly affected xylanase production. NaNO3, maltose and KCl showed significant effects on xylanase production. These three medium components were further optimized using a 3-level Box-Behnken design, and their optimum levels were maltose, 120 g/L; NaNO3, 12 g/L and KCl, 2 g/L. A xylanase activity of 1,620 U/mL, (12,460 U/g of maltose) was observed when using the optimum medium, which was 280% greater than the maximum level obtained with the basic medium. Pyridoxine concentration in the optimum medium was reduced to limit growth and divert substrate toward enzyme production. Reducing pyridoxine from 1,000 μg/ml to 300 μg/ml increased xylanase activity to 1,790 U/ml. Optimized medium with reduced pyridoxine was then tested in a stirred tank bioreactor, which resulted in 112% more xylanase activity and 43% less dry cell weight compared with the medium containing 1,000 μg/ml of pirydoxine.

Effect of trophic, osmotically active and solidifying components of nutrient medium on the direct embryognesis in spring barley anther culture in vitro

Aim. Ambryoidogenesis, or somatic embryogenesis, is known to be the most efficient mode of plant regeneration in plant cell, tissue and organ culture. The investigation was aimed to elucidate effects of trophic and osmogenic components of inductive medium on the frequency of direct embryoidogenesis in spring barley anther culture in vitro and to determine mechanism of morphogenesis improvement coursed by chemically modified starch D-5aM used as a gelling agent instead of agar. Methods. Anthers of DH-line with a high androgenetic capacity were inoculated on inductive media containing N6 macro-, MS micronutrients, organic supplements, maltose or mannitol (0.3 M) and solidified with agar or chemically modified starch. Results. A positive effect of combination of high maltose content and chemically modified starch on the induction and regeneration processes in spring barley anther culture in vitro was confirmed. It was also shown that mannitol didn’t keep any growth or development processes going in barley anther and embryo culture, but at the same time this substance had no toxic effect. Conclusions. In order to achieve a high frequency of induction in spring barley anther culture, it is necessary to use medium containing maltose – a low weight component both with trophic and with osmotic activity. Keywords: Hordeum vulgare L., anther culture in vitro, mannitol, maltose, embryo formation, plant regeneration.

Optimal recultivation conditions of Candida Albicans staying in non-culturable state

Optimal conditions for recultivation of Candida albicans staying in viable but non-culturable state have been established. The effect of sodium hypochlorite on the transition of this culture into non-culturable state was investigated. The conditions of the culture reactivation were determined. These conditions include the effect of M-9 nutrient medium components and the cultivation temperature on recultivation process.

Optimal Production of a Rhodococcus erythropolis ATCC 4277 Biocatalyst for Biodesulfurization and Biodenitrogenation Applications

Rhodococcus sp. has a broad catabolic diversity and unique enzymatic capabilities, and it is able to adapt under extreme conditions. Thereby, the production of this remarkable bacterium has a great biotechnological and industrial importance. In this sense, we sought to improve the R. erythropolis ATCC 4277 growth through a central composite design, by varying the components of nutrient medium (glucose, malt extract, yeast extract, CaCO3), temperature, and agitation. It was found that the concentrations of glucose and malt extract are not statistically significant, being reduced of 4.0 and 10.0 g L-1 to 2.0 and 5.0gL-1, respectively. The CaCO3 concentration and temperature were also diminished of 2.0 to 1.16gL-1and 28 to 23.7°C, respectively. Optimal growth conditions provided a 240% increase in final biomass concentration, an increment in specific growth rate, and a growth yield coefficient about five times greater. Application of the optimal conditions in biodesulfurization and biodenitrogenation processes showed that desulfurization capability is not associated with optimal growth conditions; however, it was achieved a 47% of nitrogen removal in the assay containing 10% (w/w) of heavy gas oil. Graphical Abstract ᅟ.

English

Trichoderma viride Sn-9106 with high cellulase activity was used to produce enzyme on residues of Chinese herbs as substrate in solid state fermentation. Residues of Chinese herbs and peptone were found to be the best combination of carbon and nitrogen source for the production of cellulase. The nutrient composition of medium was optimized using response surface methodology. A fractional factorial design (33) was applied to elucidate the nutrient medium components that significantly affect cellulase production. The concentration of peptone and wheat bran in the medium was a significant factor. The composition of nutrient fermentation medium optimized with response surface methodology was in g/L: wheat bran, 19.8, peptone, 2.06 and KH2PO4, 2.9. Compared to the original medium, the cellulase activity increased from 3.8 to 7.5 IU/mL. Key words: cellulase, Trichoderma viride Sn-9106, response surface methodology (RSM), solid state fermentation (SSF), residues of Chinese herbs (RCH).

Optimization of the fermentation medium for Paecilomyces tenuipes N45 using statistical approach

In the present study, a sequential statistical approach was applied to optimize the medium in submerged cultivation of Paecilomyces tenuipes N45. Desirability value was used as response to simultaneously enhance the yields of mycelium, adenosine, polysaccharide and cordyceps acid. Based on single-factor optimization strategy, the suitable carbon sources, nitrogen sources and inorganic salts were obtained. Then key medium components were identified by Plackett-Burman design (PBD) and further optimized by Box-Behnken design (BBD). Finally, response surface methodology (RSM) and artificial neural network – genetic algorithm (ANN-GA) were used to model and optimize the experimental results obtained from BBD. The optimum components of nutrient medium comprised (g/L): glucose 40, beef extract 10, soy peptone 10, KH2PO4 0.688, MgSO4·7H2O 1, NaCl 0.500, VB1 0.201, VB12 0.130. In a word, a mean value of desirability valuesDv = 0.493 was obtained, which was 20.540% higher than the value achieved by the basal medium. The biomass, the production of adenosine, the polysaccharide and the cordyceps acid yields were enhanced by 8.200, 3.580, 23.170 and 31.510% respectively. Key words: Artificial neural network, optimization, Paecilomyces tenuipes, response surface methodology, genetic algorithm.

High efficiency canthaxanthin production by a novel mutant isolated from Dietzia natronolimnaea HS-1 using central composite design analysis

The bacterium Dietzia natronolimnaea HS-1 is biotechnologically important due to its ability to biosynthesis the pigment canthaxanthin (CX). This study presents a promising direction to attain to the highest feasible levels in biomass dry weight (BDW), total carotenoid (TC) and CX by using the selection of superior mutant type in combination with optimization of nutrient medium components. The mutant strains were obtained using UV irradiation and ethyl methane sulfonate (EMS) treatment. Results showed that the mutants induced by EMS had higher survival and growth rates compared to UV mutants. EMS was effective mutagenic agent for strain improvement of D. natronolimnaea HS-1 for enhanced CX productivity. Also, preliminary batch experiments were carried out to achieve the suitable conditions for CX production analysis and found that d-glucose, mannose and Fe3+ have significant influences on the CX biosynthesis and microorganism growth. The medium formulation in a batch bioreactor was optimized by a central composite design-response surface methodology (CCD-RSM). Second-order empirical equations with high R2 (0.989–0.996) between the responses and independent variables were derived. The optimum conditions for the maximum of BDW (9.95g/l), TC (7.67mg/l) and CX (7.10mg/l) amounts were: d-glucose (25g/l), mannose content (15.12g/l) and Fe3+ concentration (36.77ppm).

Вплив складу живильного середовища на розвиток та проростання незрілих зародків кукурудзи в культурі in vitrо

The effect of mineral components of nutrient medium and abscisic acid (ABA) on isolated immature maize embryos is investigated with the aim to intensify their growth and to inhibit their early germination. Abscisic acid together with MS medium leads to the stasis of the development of both seedlings and embryos. The application of ABA along withN6 minerals inhibited the development of seedling’s organs in favour of embryos growth. For production of seedlings from immature embryos of 1 mm long their cultivation on modified N6 medium with addition of 12 mg/Lof abscisic acid is recommended.

On possible reasons for the selective heteroadagulation of mesenchyme human cells to the surface of acrylic hydrogels

Changes in the electrokinetic potential and hydrophilicity of hydrogels with different compositions occurring under the effect of adsorption of the components of nutrient media (initial and after 5 days of contact with mesenchyme 4BL2 human cells) are studied. It is shown that the selectivity of cell heteroadagulation to such hydrogels can be caused by the selective adsorption of water-soluble cell exometabolites on hydrogel surfaces.

Optimization of nutrient medium containing agricultural wastes for xylanase production by Aspergillus niger B03 using optimal composite experimental design

The xylanase biosynthesis is induced by its substrate – xylan. The high xylan content in some of the wastes like corn cobs and wheat bran makes them an accessible and cheap source of inducers. Nutrient medium for xylanase biosynthesis in submerged cultivation of Aspergillus niger B03 has been optimized. The optimization process was analyzed using optimal composite experimental design and response surface methodology. The predicted by the regression model optimum components of nutrient medium are as follows (g/l): (NH 4) 2HPO 4 2.6, urea 0.9, corn cobs 24.0, wheat bran 14.6 and malt sprout 6.0. Five parallel experiments have been carried out, at definite, optimum components concentrations of the nutrient medium, and a mean value of the activity Y = 996.30 U/ml has been obtained. The xylanase activity, obtained with the optimized nutrient medium is 33% higher than the activity, achieved with the basic medium.