Establishment of efficient system in the DNA microinjection into porcine in vitro embryos

2014 ◽  
Author(s):  
Don Buddika Oshadi Malaweera ◽  
Sisitha Ramachandra ◽  
Ghangyong Kim ◽  
Seokjoong Kim ◽  
Jongki Cho
Keyword(s):  
Efficient System ◽  
Dna Microinjection

scholarly journals In Vitro Co-Exposure to CeO2 Nanomaterials from Diesel Engine Exhaust and Benzo(a)Pyrene Induces Additive DNA Damage in Sperm and Cumulus Cells but Not in Oocytes

Nanomaterials ◽  
2021 ◽  
Vol 11 (2) ◽  
pp. 478
Author(s):  
Martina Cotena ◽  
Mélanie Auffan ◽  
Virginie Tassistro ◽  
Noémie Resseguier ◽  
Jérôme Rose ◽  
...  
Keyword(s):  
Dna Damage ◽  
Diesel Engine ◽  
Fossil Fuels ◽  
Cumulus Cells ◽  
Ambient Air ◽  
Cell Types ◽  
Reproductive Organs ◽  
Efficient System ◽  
Polycyclic Aromatic

Benzo(a)pyrene (BaP) is a recognized reprotoxic compound and the most widely investigated polycyclic aromatic hydrocarbon in ambient air; it is widespread by the incomplete combustion of fossil fuels along with cerium dioxide nanomaterials (CeO2 NMs), which are used in nano-based diesel additives to decrease the emission of toxic compounds and to increase fuel economy. The toxicity of CeO2 NMs on reproductive organs and cells has also been shown. However, the effect of the combined interactions of BaP and CeO2 NMs on reproduction has not been investigated. Herein, human and rat gametes were exposed in vitro to combusted CeO2 NMs or BaP or CeO2 NMs and BaP in combination. CeO2 NMs were burned at 850 °C prior to mimicking their release after combustion in a diesel engine. We demonstrated significantly higher amounts of DNA damage after exposure to combusted CeO2 NMs (1 µg·L−1) or BaP (1.13 µmol·L−1) in all cell types considered compared to unexposed cells. Co-exposure to the CeO2 NMs-BaP mixture induced additive DNA damage in sperm and cumulus cells, whereas no additive effect was observed in rat oocytes. This result could be related to the structural protection of the oocyte by cumulus cells and to the oocyte’s efficient system to repair DNA damage compared to that of cumulus and sperm cells.

scholarly journals Nature, Position, and Frequency of Mutations Made in a Single Cycle of HIV-1 Replication

2010 ◽  
Vol 84 (19) ◽  
pp. 9864-9878 ◽  
Author(s):  
Michael E. Abram ◽  
Andrea L. Ferris ◽  
Wei Shao ◽  
W. Gregory Alvord ◽  
Stephen H. Hughes
Keyword(s):  
Viral Replication ◽  
Mutation Rate ◽  
Hot Spots ◽  
High Rate ◽  
Published Data ◽  
Single Cycle ◽  
Efficient System ◽  
Hiv 1

ABSTRACT There is considerable HIV-1 variation in patients. The extent of the variation is due to the high rate of viral replication, the high viral load, and the errors made during viral replication. Mutations can arise from errors made either by host DNA-dependent RNA polymerase II or by HIV-1 reverse transcriptase (RT), but the relative contributions of these two enzymes to the mutation rate are unknown. In addition, mutations in RT can affect its fidelity, but the effect of mutations in RT on the nature of the mutations that arise in vivo is poorly understood. We have developed an efficient system, based on existing technology, to analyze the mutations that arise in an HIV-1 vector in a single cycle of replication. A lacZα reporter gene is used to identify viral DNAs that contain mutations which are analyzed by DNA sequencing. The forward mutation rate in this system is 1.4 × 10−5 mutations/bp/cycle, equivalent to the retroviral average. This rate is about 3-fold lower than previously reported for HIV-1 in vivo and is much lower than what has been reported for purified HIV-1 RT in vitro. Although the mutation rate was not affected by the orientation of lacZα, the sites favored for mutations (hot spots) in lacZα depended on which strand of lacZα was present in the viral RNA. The pattern of hot spots seen in lacZα in vivo did not match any of the published data obtained when purified RT was used to copy lacZα in vitro.

A “paper bridge” system to improve in-vitro propagation of arbuscular mycorrhizal fungi

Botany ◽  
2010 ◽  
Vol 88 (6) ◽  
pp. 617-620 ◽  
Author(s):  
Yolande Dalpé ◽  
Sylvie Seguin
Keyword(s):  
Arbuscular Mycorrhizal Fungi ◽  
Mycorrhizal Fungi ◽  
Arbuscular Mycorrhizal ◽  
Petri Dish ◽  
Efficient System ◽  
Arbuscular Mycorrhizal Fungal ◽  
Slow Growing ◽  
Paper Bridge ◽  
Bridge System

The in-vitro culture of arbuscular mycorrhizal fungi on excised roots, especially when performed on bi-compartmented Petri dishes, has proven to be an efficient system for the production of root-free fungal material. However, even after the contact between fungal hyphae and the excised roots in the proximal root compartment has occurred, up to several weeks may be required for the fungal runner hyphae to cross the median Petri dish wall and reach the distal fungal compartment. This delay is particularly long for the cultivation of slow-growing strains that usually colonize the substrate less aggressively. The delay is due to the difficulty the runner hyphae have in crossing the median Petri dish wall that separates compartments. To facilitate the passage of the fungus across the median wall, a “paper bridge” system has been devised and tested with a number of arbuscular mycorrhizal fungal strains. This method substantially accelerated fungal propagation and simplified the manipulations necessary. The proposed paper-bridge system is described and its advantages discussed.

scholarly journals ESTABLECIMIENTO DEL SISTEMA DE REGENERACION POR EMBRIOGÉNESIS SOMÁTICA DE Azadirachta indica A. Juss.

2014 ◽  
Vol 20 (2) ◽  
Author(s):  
Maria D. Artigas R ◽  
Rafael Fernandez Da Silva
Keyword(s):  
Somatic Embryogenesis ◽  
Azadirachta Indica ◽  
Culture System ◽  
Plant Metabolites ◽  
Secondary Somatic Embryogenesis ◽  
Short Term ◽  
Efficient System ◽  
Low Concentrations ◽  
Whole Plants

<p><em>Azadirachta indica</em>, es una planta con múltiples aplicaciones tanto forestal como farmacológica. Por ende, el establecimiento del sistema de cultivo <em>in vitro</em> por embriogénesis somática ofrece diversas y variadas ventajas, tales como obtener plantas altamente productivas en metabolitos. En este estudio, se utilizaron secciones foliares y cotiledonares, inducidas en medios MS (1962) suplementados con: BAP sólo y combinado con ANA/2,4-D, TDZ sólo y con ABA. La regeneración fue con MS sólo o con K + AIA y BAP + AIA. Como resultado se estableció un sistema eficiente con secciones de cotiledones, observándose organogénesis a bajas concentraciones de BAP, mientras a altos niveles de BAP (2,5 mg.L<sup>-1</sup>), así como  con TDZ + ABA (0,02 + 1mg.L<sup>-1</sup>) respectivamente favorecieron la embriogénesis somática primaria y secundaria en un 96 % y 71 % respectivamente. La regeneración fue 71 % con MS, mientras que el enraizamiento fue de 86,67 % con MS½, obteniéndose plantas completas a corto plazo. </p><p><strong>ABSTRACT</strong></p><p><em>Azadirachta indica</em>, is a plant with multiple forest and pharmacological application. Therefore, the establishment of <em>in vitro</em> culture system for somatic embryogenesis offers several distinct advantages such as obtaining highly productive plant metabolites. In this study, were used sections cotyledon and leaf, induced on MS medium (1962) supplemented with: BAP alone and combined with NAA / 2,4-D, TDZ alone and ABA. Regeneration was with MS alone or with K + BAP + IAA and IAA. As a result was established an efficient system with cotyledon sections, being observed organogenesis at low concentrations of BAP, while high levels of BAP (2.5 mg.L<sup>-1</sup>) with 96 % and TDZ + ABA (0.02 + 1mg.L<sup>-1</sup>) with 71 %, favoring the primary and secondary somatic embryogenesis. Regeneration was 71 % with MS; rooting was 86.67 % with MS½, presenting whole plants obtained short term.</p><p><strong>RESUMO</strong></p><p><em>Azadirachta indica, </em>é uma planta com várias aplicaçãos florestas e farmacológicas. Por conseguinte, a criação de sistema de cultur<em>a in vitro </em>para a embriogénese somática apresenta várias vantagens, tais como a obtenção de plantas altamente produtivas de metabolitos. Neste estudo, as seções de cotilédones e folhas, induzidas em meios suplementado en MS (1962) con: BAP sozinho e combinado com ANA / 2,4-D, TDZ e ABA sozinho. A regeneração foi apenas com MS ou com K + BAP + IAA e IAA. Como resultado, foi eficiente con seções cotiledones, observados organogênese em baixas concentrações de BAP, en quanto altos níveis de BAP (2,5 mg L-1) e com TDZ + ABA (0,02 + 1mg.L-1) favoreceu a embriogênese somática primária e secundária até 96 % e 71%, respectivamente. Regeneração estava com MS 71 %, en quanto que o enraizamento foi de 86,67 %, com ½ MS produzindo plantas inteiras curto prazo.</p><p><strong><br /></strong></p><p> </p>

scholarly journals Adventitious Shoot Regeneration from In Vitro Leaf Explants of the Peach Rootstock Hansen 536

Plants ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 755
Author(s):  
Angela Ricci ◽  
Luca Capriotti ◽  
Bruno Mezzetti ◽  
Oriano Navacchi ◽  
Silvia Sabbadini
Keyword(s):  
In Vitro Regeneration ◽  
Adventitious Shoots ◽  
Leaf Explants ◽  
Basal Medium ◽  
Adventitious Shoot ◽  
Shoot Cultures ◽  
Efficient System ◽  
Regeneration Rate ◽  
Factors Affecting

In the present study, an efficient system for the in vitro regeneration of adventitious shoots from the peach rootstock Hansen 536 leaves has been established. Twenty regeneration media containing McCown Woody Plant Medium (WPM) as a basal salt supplemented with different concentrations and combinations of plant growth regulators (PGRs) were tested. Expanded leaves along with their petiole from 3-week-old elongated in vitro shoot cultures were used as starting explants. The highest regeneration rate (up to 53%) was obtained on WPM basal medium enriched with 15.5 μM N6-benzylaminopurine (BAP). The influences on leaf regeneration of the ethylene inhibitor silver thiosulphate (STS) and of different combinations of antibiotics added to the optimized regeneration medium were also investigated. The use of 10 μM STS or carbenicillin (238 μM) combined with cefotaxime (210 μM) significantly increased the average number of regenerating shoots per leaf compared to the control. In vitro shoots were finally elongated, rooted and successfully acclimatized in the greenhouse. The results achieved in this study advances the knowledge on factors affecting leaf organogenesis in Prunus spp., and the regeneration protocol described looks promising for the optimization of new genetic transformation procedures in Hansen 536 and other peach rootstocks and cultivars.

Long-term In Vitro Toxicity Models: Comparisons Between a Flow-cell Bioreactor, a Static-cell Bioreactor and Static Cell Cultures

2002 ◽  
Vol 30 (5) ◽  
pp. 515-523 ◽  
Author(s):  
Patricia Pazos ◽  
Salvador Fortaner ◽  
Pilar Prieto
Keyword(s):  
Cell Culture ◽  
Cell Viability ◽  
Flow Cell ◽  
Model Systems ◽  
In Vitro Toxicity ◽  
Protein Determination ◽  
Efficient System ◽  
Cytotoxicity Studies

In vitro long-term toxicity testing is becoming an important issue in the field of toxicology, and there is a need to develop new model systems that mimic human chronic exposure and its effects. The aim of this work was to test two long-term in vitro toxicity systems which are available, a flow-cell bioreactor (Tecnomouse) and a static cell bioreactor system (CELLine CL 6-well), and to compare them with the use of conventional cell culture flasks. A human cell line, Int 407, was exposed to cadmium chloride (CdCl2; 10–7–10–8M) for 4 weeks. Cell numbers and cell viabilities were determined by the trypan blue (TB) exclusion assay and from exclusion of propidium iodide (PI) as determined by flow cytometry; and cell viability and metabolic activity were determined by the MTT assay. In addition, total protein determination and cadmium uptake measurements were performed. The results obtained with TB and PI exclusion did not show clear differences in cell viability with increasing CdCl2 concentration. However, in the static cell-culture systems, an increase in MTT reduction was found at low concentrations of CdCl2. Expression of heat-shock protein (Hsp27 and Hsp70) increased differently, depending on the CdCl2 concentration applied and the system used. In summary, of the two bioreactors, the CELLine CL 6-well bioreactor was shown to be the more efficient system for performing long-term cytotoxicity studies. It is easy to handle, it permits the assessment of several endpoints, and sufficient replicates can be made available.

scholarly journals In vitro Regeneration and Callogenesis of Libidibia ferrea

2020 ◽  
pp. 14-24
Author(s):  
Daniel da Silva ◽  
Angela Maria Imakawa ◽  
Kamylla Rosas Vieira Guedes ◽  
Flávio Mauro Souza Bruno ◽  
Paulo de Tarso Barbosa Sampaio
Keyword(s):  
Culture Media ◽  
In Vitro Regeneration ◽  
Shoot Multiplication ◽  
Light Sources ◽  
Multiplication Rate ◽  
Medicinal Species ◽  
Efficient System ◽  
Blue Led ◽  
Friable Callus

Libidibia ferrea (Fabaceae) is a valuable medicinal species in the Amazon, but as it is a protected plant, collection from natural populations is forbidden. Therefore, establishing an efficient system for in vitro regeneration and to improve callogenesis of this species is desirable. To determine the optimal nutritional factors needed for shoot multiplication and callus induction, different culture media, plant growth regulators and LED light sources were tested. The data were subjected to analysis of variance (ANOVA) and means compared by Tukey’s test at p < 0.05. We observe that explants inoculated in the Murashige and Skoog (MS) media with 0.05 mg L-1 of 6-benzilaminopurine (BAP) and cultivated under red-blue LED induced the highest number of shoots (3.67), number of buds (3.13), multiplication rate (15.67) and shoots length (22.03 mm) when compared with other treatments. MS and B5 media supplemented with 2.21 and 4.42 mg L-1 of 2,4-D induced 100% formation of friable callus cultivated under red-blue LED, demonstrating that the light quality significantly influenced callogenesis. Obtained results confirmed that in vitro regeneration and callogenesis is a useful strategy in the protection of endangered species. In this way, a new renewable source of biomass with high quality plant material is presented aiming at the bioprospecting of seedling extracts and friable callus to obtain secondary metabolites of this medicinal plant.

scholarly journals Molecular Identification of Endophytic Fungi and Their Pathogenicity Evaluation Against Dendrobium nobile and Dendrobium officinale

2020 ◽  
Vol 21 (1) ◽  
pp. 316 ◽  
Author(s):  
Surendra Sarsaiya ◽  
Archana Jain ◽  
Qi Jia ◽  
Xiaokuan Fan ◽  
Fuxing Shu ◽  
...  
Keyword(s):  
Molecular Identification ◽  
Endophytic Fungi ◽  
Fungal Endophytes ◽  
Dendrobium Officinale ◽  
Plant Host ◽  
Efficient System ◽  
Dendrobium Nobile ◽  
Living Tissues ◽  
Pathogenic Effects

Dendrobium are tropical orchid plants that host diverse endophytic fungi. The role of these fungi is not currently well understood in Dendrobium plants. We morphologically and molecularly identified these fungal endophytes, and created an efficient system for evaluating the pathogenicity and symptoms of endophytic fungi on Dendrobium nobile and Dendrobium officinale though in vitro co-culturing. ReThe colony morphological traits of Dendrobium myco-endophytes (DMEs) were recorded for their identification. Molecular identification revealed the presence of Colletotrichum tropicicola, Fusarium keratoplasticum, Fusarium oxysporum, Fusarium solani, and Trichoderma longibrachiatum. The pathogenicity results revealed that T. longibrachiatum produced the least pathogenic effects against D. nobile protocorms. In seedlings, T. longibrachiatum showed the least pathogenic effects against D. officinale seedlings after seven days. C. tropicicola produced highly pathogenic effects against both Dendrobium seedlings. The results of histological examination of infected tissues revealed that F. keratoplasticum and T. longibrachiatum fulfill Koch’s postulates for the existence of endophytes inside the living tissues. The DMEs are cross-transmitted inside the host plant cells, playing an important role in plant host development, resistance, and alkaloids stimulation.

Optimizing proliferation and assessment of valak morpho-phenological traits; an endangered nutritious Allium endemic to Iran

2021 ◽  
Author(s):  
Sajad Jafari ◽  
Mohammadreza Hassandokht ◽  
Madi Taheri
Keyword(s):  
Genetic Resources ◽  
Dry Weight ◽  
Growth Cycle ◽  
Phenological Stages ◽  
Efficient System ◽  
Seedling Length ◽  
Conservation Of Genetic Resources ◽  
Medicinal Properties ◽  
One Year

Abstract Allium elburzense W. (subg. Melanocrommyum ) is an endangered and endemic species to Iran, which is called “valak” and known as a valuable and pricy vegetable with high nutritional and medicinal properties. This study was carried out in order to domestication, removing barriers of sexual reproduction, micropropagation and investigation of phenological stages and growth cycle of the species. The results indicated that fresh seeds had better germination than one year old seeds and appropriate germination temperature in A. elburzense W. seed was 12 °C. According to ANOVA results, the triple interaction of scarification (Sc), stratification period (StP) and GA 3 had a significant effect on seedling length, seedling fresh and dry weight, bulb height and diameter (p<0.01). Comparison of means showed that the treatments in which the StP (45 days) and scarification was applied had better effects on seed germination (66.5% germination) than other treatments. Murashige and Skoog with 300 mg L -1 KH2PO4 (MSP) medium caused a significant increase in the diameter of the basal plate compared to MS medium. Efficient system for in vitro propagation and conservation of valak genetic resources were the use of MSP culture medium supplemented with 0.6 mg l -1 NAA, 4 mg l -1 BA and 120 g l -1 of sucrose, which resulted in a direct propagation coefficient of 26.83 seedlings and bulblets at 18 weeks. As regards its life cycle, the juvenile phase in a low proportion of individuals (13.33%) lasted 2 years from seeding, while 87.66% of individuals were capable of producing inflorescence in the third year. Results showed that this plant is a metamorphosis and cross pollinator species. Results of present study can be used in domestication, conservation of genetic resources and proper exploitation of valak species.

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