Evaluation of different antioxidants during in vitro establishment of allspice (Pimenta dioica L. Merrill): a recalcitrant species

Objective: To evaluate the effect of different antioxidant agents during in vitro establishment of allspice (Pimenta dioica L. Merrill). Design/methodology/approach: The effect of different antioxidant agents (Methylene blue, L-cysteine, and silver nanoparticles [AnNPs]) added to Murashige and Skoog culture medium at different concentrations were studied during axenic establishment of P. dioica. A completely randomized experimental design was used. All trials were performed in triplicate. The percentage of survival, oxidation, contamination was determined, the phenols content, antioxidant capacity and lipid peroxidation. Results: The highest survival occurred with the addition of L-cysteine. The lowest percentages oxidation were observed in explants treated with L-cysteine. Treatments with 100 and 200 mg L-1 AgNPs had the lowest contamination values. L-cysteine and 50 and 100 mg L-1 AgNPs resulted in an increase in the content of soluble phenols. The highest contents of cell wall-linked phenols were obtained in treatments with 200 mg L-1 methylene blue, L-cysteine, and 200 mg L-1 AgNPs. In this study, all treatments had a reaction of scavenging/reduction mechanisms free radicals. The highest content of malondialdehydes was observed in the control treatment and 200 mg L-1 methylene blue. The highest content of malondialdehydes was observed in the control treatment and 200 mg L-1 methylene blue. Limitations on study/implications: The highest percentage of oxidation was observed in the control treatments, 100 and 200 mg L-1 methylene blue, causing cell death. Findings/conclusions: The addition of L-cysteine to the culture medium is alternative to reduce oxidation during in vitro introduction of P. dioica.

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Evaluation of different antioxidants during in vitro establishment of allspice (Pimenta dioica L. Merrill): a recalcitrant species

10.21203/rs.3.rs-21190/v1 ◽

2020 ◽

Author(s):

Sugey Vásquez-Hernández ◽

Carlos Alberto Cruz-Cruz ◽

Maricela Santiago-Santiago ◽

Jericó Jabín Bello-Bello

Keyword(s):

Lipid Peroxidation ◽

Methylene Blue ◽

Cell Wall ◽

Antioxidant Capacity ◽

Culture Medium ◽

Control Treatment ◽

Soluble Phenols ◽

In Vitro Establishment ◽

Pimenta Dioica

Abstract Background: Pimenta dioica L. Merrill is a tree whose fruits are used as a spice due to their culinary and therapeutic uses. Conventional propagation techniques using seeds and cuttings do not guarantee the phytosanitary quality of this crop. Therefore, the use of Plant Tissue Culture techniques are an option for in vitro establishment. The aim of this study was to evaluate the effect of different antioxidant agents (Methylene blue, L-cysteine and silver nanoparticles) added to MS (Murashige and Skoog) culture medium at different concentrations (0, 50, 100 and 200 mg L-1) during axenic establishment of buds used as explants of P. dioica. Results: The percentage of survival, oxidation and contamination was determined, as well as the content of soluble phenols, cell wall-linked phenols, antioxidant capacity and lipid peroxidation. Results showed significant differences among the different antioxidants for the evaluated variables; the highest survival occurred in the treatments with the addition of L-cysteine with percentages greater than 40 %, while the lowest survival occurred in the control treatment, 100 and 200 mg L-1 methylene blue, with 0, 3.3 and 0% survival, respectively. The highest percentage of oxidation was observed in the control treatments, 100 and 200 mg L-1 methylene blue with 96.67% oxidation, while the lowest percentages were observed in explants treated with L-cysteine, with 30% oxidation. Treatments with 100 and 200 mg L-1 AgNPs had the lowest contamination values, with 20%. Biochemical determinations showed that L-cysteine and 50 and 100 mgL-1 AgNPs resulted in an increase in the content of soluble phenols. The highest contents of cell wall-linked phenols were obtained in treatments with 200 mg L-1 methylene blue, L-cysteine, and 200 mg L-1 AgNPs. Analysis of antioxidant capacity revealed that all treatments had a reaction of scavenging / reduction mechanisms free radicals. Regarding lipid peroxidation, the highest content of malondialdehydes was observed in the control treatment and 200 mgL-1 methylene blue. Conclusion: the addition of L-cysteine to the culture medium showed a higher survival rate, decreased oxidation, greater production of phenolic compounds, increased antioxidant capacity and decreased lipid peroxidation, this amino acid being an alternative to reduce oxidation during in vitro introduction of allspice and other species that exhibit recalcitrance in vitro during establishment.

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Clonal bamboo production based on in vitro culture

Bioscience Journal ◽

10.14393/bj-v36n4a2020-48169 ◽

2020 ◽

Vol 36 (4) ◽

Author(s):

Anatálya dos Santos Ribeiro ◽

Alexssandra Jéssica Rondon de Figueiredo ◽

Gabriela Cristina Rech Tormen ◽

André Luís Lopes da Silva ◽

Wellington Ferreira Campos ◽

...

Keyword(s):

Activated Carbon ◽

Large Scale ◽

Culture Medium ◽

Clonal Plant ◽

Adventitious Rooting ◽

Clonal Plants ◽

Ex Vitro ◽

Bambusa Vulgaris ◽

In Vitro Establishment

Bamboo species are an alternative for the composition of forest plantations. However, their potential has not been explored due to the hard time in producing large-scale clonal plants. Thus, the aim this work was to evaluate the in vitro establishment, bud multiplication and ex vitro rooting of Bambusa vulgaris. The first experiment tested different systemic and contact fungicide solutions, based on exposure time, during the establishment phase. Established explants were subjected to evaluation of residual fungicide effect on subcultures during the multiplication and elongation phases. The second experiment evaluated the influence of activated carbon on ex vitro survival and on adventitious rooting. Explant immersion in liquid culture medium added with 1.0 mL of fungicide for 120 hours has favored the in vitro establishment and reduced fungal contamination. On the other hand, it favored the shoot emission of shoots per explant during the multiplication phase. Both rooting induction culture medium and mini-incubator system use were effective in enabling adventitious root formation. The presence of activated carbon in the rooting induction culture medium resulted in a higher clonal plant survival rate.

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Temporary immersion biorreators: efficient technique for the propagation of the ‘Pircinque’ strawberry

Revista Brasileira de Fruticultura ◽

10.1590/0100-29452019102 ◽

2019 ◽

Vol 41 (1) ◽

Author(s):

Samila Silva Camargo ◽

Leo Rufato ◽

Maicon Magro ◽

André Luiz Kulkamp de Souza

Keyword(s):

Liquid Medium ◽

Culture Medium ◽

Culture Media ◽

Time Factors ◽

Efficient Technique ◽

Control Treatment ◽

Solid Culture ◽

Temporary Immersion ◽

Solid Culture Medium

Abstract The in vitro propagation technique via temporary immersion bioreactors is a tool that, through the culture in a liquid medium, allows an increase in the efficiency of seedling production. Several researches with the strawberry crop have shown greater efficiency of the system compared to the conventional process of micropropagation in solid medium. In this sense, the objective herein was to establish a protocol of multiplication and rooting of the ‘Pircinque’ strawberry, in temporary immersion bioreactors. Two distinct and independent studies were carried out, characterized by the multiplication and rooting stages of strawberry explants, newly introduced and registered in Brazil. Two culture media (MS and KNOP) were studied and, as a control treatment, the growth of the explants in solid culture medium was evaluated with the addition of 5 g L-1 of agar. Different immersion times of the culture medium were explored: five or eight times a day, for 15 minutes. The study was composed of the culture medium and immersion time factors, as well as the control (solid) treatment. It was verified that the use of temporary immersion bioreactors system is an efficient technique for the multiplication and rooting of explants of strawberry cv. Pircinque, when compared to the conventional method of micropropagation with the use of solid culture medium, making it possible to optimize the production of seedlings in biofactories. The MS liquid medium, in contact with explants of ‘Pircinque’ strawberry five times a day, increased the growth of the aerial part and the root system.

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In vitro establishment of Bambusa oldhamii Munro from field-grown matrices and molecular identification of endophytic bactéria

Pesquisa Agropecuária Tropical ◽

10.1590/1983-40632019v4953673 ◽

2019 ◽

Vol 49 ◽

Author(s):

Ana Paula de Azevedo Pasqualini ◽

Gabriela Xavier Schneider ◽

Hugo Pacheco de Freitas Fraga ◽

Luiz Antonio Biasi ◽

Marguerite Quoirin

Keyword(s):

Molecular Identification ◽

Culture Medium ◽

Bacterial Isolate ◽

Fungal Growth ◽

Endophytic Bacterium ◽

Liquid Culture Medium ◽

Rdna Sequencing ◽

In Vitro Establishment ◽

Bambusa Oldhamii

ABSTRACT In plant micropropagation, the establishment stage is difficult, due to the presence of microorganisms in tissues from field-grown matrices, especially for bamboo. This study aimed to establish an efficient asepsis protocol for Bambusa oldhamii explants from field plants, as well as to carry out the molecular identification of a possible endophytic bacterial isolate. The explants were exposed to 70 % alcohol, 1 % sodium hypochlorite (NaOCl), 0.1 % mercuric chloride (HgCl2), thiophanate-methyl (Cercobin®) and chlorhexidine digluconate (2 % Riohex®) in different combinations, and introduced into Murashige and Skoog culture medium (solid or liquid), supplemented or not with 4 mL L-1 of Plant Preservative Mixture (PPMTM), totaling seven treatments. The asepsis and immersion of the explants in the liquid culture medium containing 4 mL L-1 of PPMTM visually inhibited the bacterial and fungal growth, allowed the development of shoots with a mean length of 2.2 cm and posterior subcultures, being the best treatment used for the in vitro establishment of B. oldhamii. The molecular identification of an endophytic bacterium performed by 16S rDNA sequencing allowed to identify the bacterial isolate as Ralstonia sp., with 100 % of similarity, and the phylogenetic analysis grouped it with Ralstonia pickettii. In addition, the bacterial isolate showed to be sensitive to 4 mL L-1 of PPMTM by the minimum inhibitory concentration test.

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Effects of a commercial biocide, kasugamycin and consistency of the culture medium on the in vitro establishment of bamboo1

Pesquisa Agropecuária Tropical ◽

10.1590/1983-40632019v4955435 ◽

2019 ◽

Vol 49 ◽

Author(s):

Daniela Werner Ribeiro dos Santos ◽

Théo Piucco Rocker ◽

Thiago Sanches Ornellas ◽

Miguel Pedro Guerra

Keyword(s):

Liquid Culture ◽

Culture Medium ◽

Liquid Culture Medium ◽

Bambusa Vulgaris ◽

Phyllostachys Bambusoides ◽

In Vitro Establishment ◽

Dendrocalamus Asper

ABSTRACT The contamination by microorganisms and oxidation of explants in the in vitro establishment of bamboo are recurrent problems for its micropropagation. In the present study, effects of the biocide Plant Preservative Mixture (PPM™), the antibiotic kasugamycin and the consistency of the culture medium were evaluated in the in vitro establishment of Bambusa vulgaris,Phyllostachys bambusoides and Dendrocalamus asper. The presence of PPM™ in the culture medium had a significant effect using 2 mL L-1 or 4 mL L-1 concentrations, as well as in the liquid culture medium, increasing the plants established in the autumn. Kasugamycin promoted variable responses for all the three species, depending on the season. There was interaction among the factors, demonstrating that higher rates of viable plants can be obtained by combining different strategies to reduce the oxidation and contamination. For the in vitro establishment of B. vulgaris,P. bambusoides and D. asper, it is recommended to add 2 mL L-1 or 4 mL L-1 of PPM™ to the liquid culture medium.

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Polpa de banana e sacarose no desenvolvimento in vitro de orquídeas

Revista Agraria Academica ◽

10.32406/v4n2/2021/70-77/agrariacad ◽

2021 ◽

Vol 4 (2) ◽

pp. 70-77

Author(s):

Eliane Lima de = Aquino ◽

◽

Tarcísio Rangel do Couto ◽

João Sebastião de Paula Araújo ◽

◽

...

Keyword(s):

Experimental Design ◽

Seedling Growth ◽

Culture Medium ◽

In Vitro Cultivation ◽

Fresh Weight ◽

Randomized Experimental Design ◽

Number Of Leaves ◽

Weight Number

The objetive of this study was to evaluate the effects of adding two types of banana pulp, combined with varying concentrations of sacarose on the growth of Cattleya sp. plantlets. Hybrid LCTV-01 seedlings (Cattleya labiata rubra x Cattleya labiata semi alba) made to germinate in vitro were inoculated in an MS culture medium with half the concentration of nutrients and supplemented with 60 g.L-1 'maçã' or 'terra' banana pulp in addition to different concentrations of sacarose (10, 20 and 30 g.L-1. The entirely randomized experimental design was chosen, implemented in seven treatments, ten repetitions and eight seedlings per repetition. After 160 days of in vitro cultivation, variables of fresh weight, number of leaves, number of roots and length of the longest root were evaluated. It was found that the addition of banana pulp of any of the analyzed cultivars promoted better seedling growth. Additionally, the 20 g.L-1 sacarose concentration yielded better results for the analyzed variables.

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EFFECT OF 6- BENZYLAMINOPURINE AND THE CULTURE MEDIUM MS (1962) ON THE IN VITRO ESTABLISHMENT OF Prosopis pallida (WILLD.) KUNTH

REBIOL ◽

10.17268/rebiol.2019.39.02.03 ◽

2019 ◽

Vol 39 (2) ◽

pp. 30-40

Author(s):

Diego Campos ◽

Claudia Chávez ◽

Segundo Lopéz ◽

Armando Gil- Rivero ◽

Angélica Lopéz -Zavaleta ◽

...

Keyword(s):

Culture Medium ◽

In Vitro Establishment

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149 EFFECT OF RESVERATROL ANALOGUE ON DEVELOPMENT OF IN VITRO-FERTILIZED BOVINE EMBRYOS

Reproduction Fertility and Development ◽

10.1071/rdv29n1ab149 ◽

2017 ◽

Vol 29 (1) ◽

pp. 183 ◽

Cited By ~ 1

Author(s):

T. A. Patrocínio ◽

C. A. C. Fernandes ◽

L. S. Amorim ◽

J. R. Ribeiro ◽

G. C. Macedo ◽

...

Keyword(s):

In Vitro Culture ◽

Embryo Development ◽

Culture Medium ◽

Synthetic Analogue ◽

Bovine Embryo ◽

High Humidity ◽

Cleavage Rate ◽

Antioxidant Agents ◽

Blastocyst Rate

Oxidative stress is one of the main effects of in vitro culture. Generation of reactive oxygen species (ROS) by embryos can be enhanced by the sub-optimal in vitro culture conditions and are associated with a delay in embryonic development. However, supplementation of culture medium with antioxidant agents can minimize the effects of ROS (Guérin et al. 2001 Hum. Reprod. Update 7, 175–189). Resveratrol is an example of a potent antioxidant, and modifications in its structure can improve its biological activity. This study evaluated the effect of AR33 (formula with patent pending), an analogue of resveratrol with high antioxidant activity, on embryo development. Bovine cumulus-oocyte complexes recovered from ovaries collected at the slaughterhouse were in vitro matured for 24 h and oocytes were in vitro fertilized for 20 h, both at 38.8°C under 5% CO2 in air and high humidity. Partially denuded presumptive zygotes were randomly distributed in 4 treatments (with 6 replicates): 0 µM (control, n = 347), 0.1 µM (n = 337), 0.5 µM (n = 277), and 2.5 µM (n = 343) of AR33. The base medium was SOFaa supplemented with 2.5% FCS and incubation conditions were 38.8°C under 5% CO2 in air and high humidity. Half of culture medium was renewed (feeding) at Day 3 and 5 post-fertilization. Cleavage was evaluated at Day 3 and blastocyst rates at Day 7 and 8 post-fertilization. Data were analysed by logistic regression considering the significance level of P < 0.05. Values are shown as mean ± SEM. Cleavage rate was higher (P < 0.05) for 2.5 µM (69.0 ± 4.4%) than for 0, 0.1, and 0.5 µM AR33 (62.1 ± 2.0%, 60.7 ± 5.9%, and 56.7 ± 5.8%, respectively). At Day 7, the blastocyst rate was similar (P > 0.05) among 0.1, 0.5, and 2.5 µM (18.1 ± 5.4%, 17.5 ± 2.9%, and 19.4 ± 3.3%, respectively) and all of them were higher (P < 0.05) than 0 µM AR33 (12.4 ± 2.5%). At Day 8, there was again no difference (P > 0.05) among 0.1, 0.5, and 2.5 µM AR33 (21.0 ± 5.0%, 18.4 ± 2.1%, and 24.6 ± 3.3%, respectively) but only 0.1 and 2.5 µM showed higher (P < 0.05) blastocyst rate than 0 µM AR33 (15.2 ± 2.5%). In conclusion, the synthetic analogue of resveratrol tested in this study can improve bovine embryo development in culture medium supplemented with 2.5% FCS under 5% CO2 in air. A concentration of 2.5 µM AR33 can be a choice for further studies. This study was supported by Fapemig, CAPES, and CNPq.

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Hyperhydricity reversal and clonal propagation of four-wing saltbush (Atriplex canescens, Chenopodiaceae) cultivated in vitro

Australian Journal of Botany ◽

10.1071/bt07116 ◽

2008 ◽

Vol 56 (4) ◽

pp. 358 ◽

Cited By ~ 3

Author(s):

Isaac Reyes-Vera ◽

Carol Potenza ◽

Jerry Barrow

Keyword(s):

Culture Medium ◽

Clonal Propagation ◽

Inorganic Nitrogen ◽

Basal Medium ◽

Control Treatment ◽

Culture Vessel ◽

Cumulative Frequency ◽

Atriplex Canescens ◽

Normal Morphology

In vitro propagated shoots of four-wing saltbush (Atriplex canescens, Pursh Nutt) showed severe symptoms of hyperhydricity. We show that the reversion of hyperhydric A. canescens shoots to normal shoots was significantly affected by the presence of inorganic nitrogen in the culture vessel. When the culture vessel was vented or when ammonium nitrate was deleted from Murashige and Skoog basal medium, rates of reversion were significantly higher. Although statistically significant differences were evident when comparing vented v. non-vented treatments for each medium, the modified culture medium with vented closures was consistently the best treatment, showing a total cumulative frequency of 39.7% reversion to normal morphology, compared with a total cumulative frequency of 7.1% observed in the control treatment. Resulting normal shoots also showed significant improvements in further manipulations, including rooting in vitro, transplantation to soil and survival in native sites.

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Comparison of Different Semi-Automated Bioreactors for In Vitro Propagation of Taro (Colocasia esculenta L. Schott)

Plants ◽

10.3390/plants10051010 ◽

2021 ◽

Vol 10 (5) ◽

pp. 1010

Author(s):

Eucario Mancilla-Álvarez ◽

Juan Antonio Pérez-Sato ◽

Rosalía Núñez-Pastrana ◽

José L. Spinoso-Castillo ◽

Jericó J. Bello-Bello

Keyword(s):

Culture Medium ◽

Solid Medium ◽

Survival Rates ◽

Shoot Multiplication ◽

Control Treatment ◽

Multiplication Rate ◽

Solid Culture ◽

Solid Culture Medium ◽

Semi Solid

Taro is important for its nutritional content, medicinal use, and bioethanol production. The aim of the present study was to compare different semi-automated bioreactors (SABs) during in vitro multiplication of C. esculenta. The SABs used were temporary immersion bioreactors (TIBs), SETIS™ bioreactors and ebb-and-flow bioreactors; semi-solid culture medium was used as a control treatment. At 30 d of culture, different developmental variables, determination of chlorophyll, stomatal content, and survival percentage during acclimatization were evaluated. SABs increased the shoot multiplication rate relative to the semi-solid medium; however, the SETIS™ bioreactor showed the highest shoot production, with 36 shoots per explant, and the highest chlorophyll content. The stomatal index was higher in the semi-solid medium compared to the SABs, while the percentage of closed stomata was higher in the SABs than in the semi-solid culture medium. The survival rate during acclimatization showed no differences among the culture systems assessed, obtaining survival rates higher than 99%. In conclusion, the SETIS™ bioreactor showed the highest multiplication rate; however, other bioreactor alternatives are available for semi-automation and cost reduction for micropropagation of C. esculenta.

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