In vitro development of isolated leaf primordia of Matteuccia struthiopteris

Primordia (P2–P6) at the shoot apex were excised and cultured on Knudson's medium for a period of 4 weeks. The majority of primordia developed as leaves. The length, mass, and morphological complexity of these leaves were related to initial primordium age and height. There was a consistent trend toward the production of shorter, lighter, and less complex leaves from the younger, smaller initial explants. A second set of experiments traced the developmental fate of isolated primordia (P1–P4) over a longer period of time (12 weeks). Various kinds of secondary development were observed including bud and root development. Bud numbers decreased with primordial age. On the other hand, the rate of root formation increased.

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28 SIMPLIFIED ACTIVATION METHOD TO IMPROVE THE IN VITRO DEVELOPMENT OF HANDMADE CLONED (HMC) PORCINE EMBRYOS

Reproduction Fertility and Development ◽

10.1071/rdv21n1ab28 ◽

2009 ◽

Vol 21 (1) ◽

pp. 114

Author(s):

Y. Du ◽

Z. Yang ◽

B. Lv ◽

L. Lin ◽

P. M. Kragh ◽

...

Keyword(s):

Cell Number ◽

Activation Method ◽

The Other ◽

Electrical Activation ◽

In Vitro Development ◽

Reconstructed Embryos ◽

Fetal Fibroblasts ◽

Group 2 ◽

Vitro Development

Delayed activation is commonly used in pig somatic cell nuclear transfer (SCNT) where electrical activation is followed by chemical activation. However, chemical incubation of several hours (up to 4 or 6) is logistically not very convenient even though handmade cloning (HMC) could improve the overall efficiency of pig cloning (Du et al. 2007 Theriogenology 68, 1104–1110). It was reported that a brief exposure of cycloheximide (CX) before electrical activation could significantly increase developmental rate and total blastocyst cell number when simultaneous activation was performed in micromanipulator-based pig cloning (Naruse et al. 2007 Theriogenology 68, 709–716). The purpose of our present work is to investigate whether such activation method is also applicable for pig HMC. Data were analyzed by t-test using SPSS (11.0, SPSS Inc., Chicago, IL, USA). After 42 h in vitro maturation, cumulus cells were removed. In vitro-cultured porcine fetal fibroblasts were used as donor cells. Cytoplast-fibroblast pairing, electrical fusion and activation of fused cytoplast-fibroblast pairs were performed as described previously (Kragh et al. 2005 Theriogenology 64, 1536–1545; Du et al. 2005 Cloning Stem Cells 7, 199–205). Three groups were compared due to different activation protocol. In Group 1 (control), reconstructed embryos were cultured in porcine zygote medium 3 (PZM3) supplemented with 4 mg mL–1 BSA, 5 μg mL–1 cytochalasin B (CB), and 10 μg mL–1 CX for 4 h. In Group 2 (CX priming), fused pairs and the other halves of cytoplasts were incubated in HEPES-buffered TCM-199 medium supplemented with 10% calf serum, 10 μg mL–1 CX for 10 min just before the second fusion or electrical activation. In Group 3 (CB + CX priming), treatment similar to Group 2 was performed except that additional 5 μg mL–1 CB was added for the 10-min incubation. Reconstructed embryos were in vitro cultured in the well of the well (WOW) system for 6 days. Blastocyst rates and total cell numbers of Day 6 blastocysts were evaluated. As illustrated in Table 1, embryos pretreated with both CB and CX gave the best results, with better blastocyst formation (53.8 ± 4.8%; mean ± SEM) and higher cell number (77.2 ± 5.4) compared to the other 2 groups. Our data suggested that CX and CB priming could be used as a solution to the long chemical incubation in porcine SCNT by HMC, making the embryos more receptive to electrical activation. Table 1.In vitro development of HMC reconstructed embryos with different activation protocols

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In Vitro Development of Resistance to Telithromycin (HMR 3647), Four Macrolides, Clindamycin, and Pristinamycin inStreptococcus pneumoniae

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.44.2.414-417.2000 ◽

2000 ◽

Vol 44 (2) ◽

pp. 414-417 ◽

Cited By ~ 70

Author(s):

Todd A. Davies ◽

Bonifacio E. Dewasse ◽

Michael R. Jacobs ◽

Peter C. Appelbaum

Keyword(s):

The Other ◽

In Vitro Development ◽

Development Of Resistance ◽

Subinhibitory Concentrations ◽

Erythromycin A ◽

Vitro Development

ABSTRACT The ability of 50 sequential subcultures in subinhibitory concentrations of telithromycin (HMR 3647), azithromycin, clarithromycin, erythromycin A, roxithromycin, clindamycin, and pristinamycin to select for resistance was studied in five macrolide-susceptible and six macrolide-resistant pneumococci containing mefE or ermB. Telithromycin selected for resistance less often than the other drugs.

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Fasciation d'extrémités caulinaires du Celosia cristata (Amarantacées) cultivées in vitro

Canadian Journal of Botany ◽

10.1139/b81-186 ◽

1981 ◽

Vol 59 (8) ◽

pp. 1367-1372 ◽

Cited By ~ 3

Author(s):

D. Driss-Ecole

Keyword(s):

Indoleacetic Acid ◽

Leaf Primordia ◽

Root Initiation ◽

Salt Mixture ◽

Shoot Apices ◽

In Vitro Development ◽

Long Period ◽

Celosia Cristata ◽

Vitro Development

In vitro development of fasciation was achieved from the excised shoot apices of Celosia cristata in a 16-h photoperiod. An 8-h photoperiod produced no development. The explants consisted of the meristematic dome with two leaf primordia. The best results were obtained with a nutrient medium composed of Murashige and Skoog salt mixture, 30 g/L sucrose, 5 g/L agar, and 1 mg/L indoleacetic acid (IAA). Isolated shoot apices developed callus at the bases of the explants and numerous leaves on one, two, or three flattened and fasciated meristems. Root initiation occurred sometimes but only after a long period of culture.

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EFEITO DE HERBICIDAS UTILIZADOS EM CANA-DEAÇÚCAR NO DESENVOLVIMENTO IN VITRO DO FUNGO ENTOMOPATOGÊNICO Metarhizium anisopliae (Metsch.) SOROKIN

Pesticidas Revista de Ecotoxicologia e Meio Ambiente ◽

10.5380/pes.v14i0.3119 ◽

2004 ◽

Vol 14 ◽

Author(s):

EDUARDO A. D. DA COSTA ◽

MARCUS B. MATALLO ◽

JOSÉ E. M. ALMEIDA ◽

ELISÂNGELA S. LOUREIRO ◽

ALEXANDRE H. SANO

Keyword(s):

Metarhizium Anisopliae ◽

Culture Media ◽

Average Diameter ◽

Potato Dextrose Agar ◽

The Other ◽

In Vitro Development ◽

Petri Dishes ◽

Equidistant Points ◽

Vitro Development

Avaliou-se a compatibilidade dos herbicidas butafenacil, metribuzim, s-metolacloro, 2,4-D, glifosato, oxassulfurom e trifloxissulfuron no desenvolvimento do fungo entomopatogênico M. anisopliae (isolado IBCB 348), empregado para controle biológico de Mahanarva fimbriolata (Hemiptera: Cercopidae) conhecida como cigarrinhada- raiz da cana. Os herbicidas foram adicionados ao meio de cultura batata dextrose ágar (BDA) em temperatura aproximada de 45°C nas doses de campo mínimas e máximas recomendadas, fixandose o volume de calda em 200 L/ha. Após a solidificação do meio em placas de Petri, o fungo foi inoculado em três pontos eqüidistantes. As placas foram incubadas em câmara climatizada avaliandose o diâmetro médio das colônias formadas, o número de conídios por colônia e a viabilidade dos conídios do fungo após 20 e 48 horas da inoculação. Apenas o herbicida oxassulfurom, na menor dose testada (30 g/ha), foi classificado como tóxico ao M. anisopliae, sendo os demais classificados em ambas as doses como muito tóxicos. Quanto à viabilidade, somente os herbicidas oxassulfurom nas doses de 30 e 60 g/ha não afetaram a germinação do fungo até 20 horas após a inoculação e o glifosato a 0,5 e 6,0 L/ha até 48 horas. Os demais herbicidas mostraram-se incompatíveis, afetando o desenvolvimento e a germinação do M. anisopliae. EFFECT OF HERBICIDES UTILIZED IN SUGAR CANE OF IN VITRO DEVELOPMENT OF ENTOMOPHATOGENIC FUNGI Metarhizium anisopliae (Metsch.) SOROKIN Abstract The compatibility of the herbicides butafenacil, metribuzin, s-metolachlor, 2,4-D, glyphosate, oxasulfuron and trifloxysulfuron in the development of entomopathogenic fungi M. anisopliae (isolate IBCB 348) were evaluated, which is employed in the biological control of Mahanarva fimbriolata (Hemiptera: Cercopidae). The herbicides were added to the culture media potato dextrose agar (PDA) in temperature of approximately 45º C in minimal and maximal recomended field doses, fixing the volume of 200 L/ha. After the solidification of the culture media in Petri dishes, the fungi was inoculated in three equidistant points. The plates were incubated in climatized chamber and the average diameter of the formed colonies were evaluated, the number of conidia per colony and the viability of the conidia after 20 and 48 hours inoculation. Only the herbicide oxasulfuron, in the lowest tested dose (30 g/ha) was classified as toxic to M. anisopliae, being the rest classified in both doses as highly toxic. For viability only the herbicides oxasulfuron in the doses of 30 and 60 g/ha didnt affect the germination of the fungi until 20 hours after inoculation and glyphosate at 0.5 and 6.0 L/ha until 48 hours. The other herbicides were imcompatible, affecting the development and the germination of M. anisopliae.

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Survival rates of mouse blastocyst vitrified in dimethylformamide based solutions associated with ethylene glicol or 1-2 propanediol

Ciência Rural ◽

10.1590/s0103-84782011005000139 ◽

2011 ◽

Vol 41 (11) ◽

pp. 1985-1990 ◽

Cited By ~ 3

Author(s):

Paula Rodriguez Villamil ◽

Felipe Ledur Ongaratto ◽

Daniela Scherer da Silva ◽

Berenice de Avila Rodrigues ◽

Jose Luiz Rodrigues

Keyword(s):

Ethylene Glycol ◽

Survival Rates ◽

The Other ◽

Developmental Competence ◽

Time Interval ◽

Equilibrium Solutions ◽

Mouse Blastocyst ◽

In Vitro Development ◽

Vitro Development

The aim of this study was to determine the effect of dimethylformamide (DF) associated with ethylene glycol (EG) or 1-2 propanediol (PROH) during vitrification, on the in vitro development of mouse blastocysts. Cryoprotectant toxicity was evaluated exposing embryos into three different equilibrium solutions (ES) composed by DF, EG or PROH mixtures (10% v/v of each) in mPBS + 0.5% PVA at different interval times (1, 3 and 10min). In a second experiment, embryos were exposed to the same ES (either 1 or 3min), following for the three respectively vitrification solutions (VS) (20% v/v of each) for 30s. After 72 hours of in vitro culture, embryo hatching and expansion rates were similar for the ES1 and ES2 equilibration solutions during the time interval of 1 or 3min. However embryos exposed for 10 min to the DF equilibration solutions, had lower survival rates than EG-PROH solution (P<0.01). Furthermore, survival rates for embryos exposed to DF-PROH (ES+VS) were lower than embryos exposed to the other solutions (P<0.01). Blastocyst vitrification was performed with the three ES+VS (for 1min and 30s, respectively), using glass micropipettes (GMP). Survival rates were lower for blastocysts vitrified with DF solutions (3%-3/108 and 17.1%-19/111) (P<0.01) than with PROH+EG vitrification solutions (69%-73/105). In conclusion, DF as a cryoprotectant into vitrification solutions have deleterious effects on the in vitro developmental competence of vitrified mouse blastocysts.

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138 THE EFFECT OF SYNTHETIC HYALURONAN, BSA AND SERUM ON IN VITRO DEVELOPMENT AND GENE TRANSCRIPTION OF BOVINE BLASTOCYSTS

Reproduction Fertility and Development ◽

10.1071/rdv17n2ab138 ◽

2005 ◽

Vol 17 (2) ◽

pp. 219

Author(s):

A. Gutiérrez-Adán ◽

H. Rodriguez-Martinez ◽

P. Beltrán Breña ◽

J. De la Fuente ◽

A.T. Palasz

Keyword(s):

Fine Structure ◽

Repeated Measures ◽

Expression Patterns ◽

Calf Serum ◽

The Other ◽

Bovine Embryo ◽

In Vitro Development ◽

Chi Square Analysis ◽

Vitro Development

The objective of this study was to examine the effect of synthetic hyaluronan (s-HA), BSA and fetal calf serum (FCS) on bovine embryo in vitro development, ultrastructure, and mRNA transcription of four developmentally important genes: apoptosis (BAX), oxidative stress (SOX), growth factor (IGF-II), and cell-to-cell adhesion (Ecad). A total of 1406 presumptive zygotes (7 replicates) were cultured initially in two Groups: 1, SOFaa + 4% BSA only, and 2, SOFaa + 4% BSA and 10% FCS. On Day 4 (96 h after insemination) of culture, the number of zygotes that developed to the <8-cell-stage were recorded, and 2.5 mg/mL of s-HA (MAP-5; Bioniche Inc, Belleville, ON, Canada) was added to half of the embryos from each group; the other half received an equivalent volume of corresponding SOF medium without s-HA. Embryos were cultured in 50-μL drops (25 zygotes per drop) under paraffin oil at 39°C and 5% CO2 in humidified air. Cleavage rates were recorded on Day 2 and the number of blastocysts on Days 7, 8, and 9. At least five blastocysts from each replicate and from each treatment were frozen for evaluation of gene expression patterns. Poly(A) mRNA was prepared from 4–5 groups of pools of 10 embryos. The quantification of all gene transcripts was performed by real time quantitative RT-PCR in three replicates. The fine structure of blastocysts was studied using transmission electron microscopy. Embryo developmental stages and blastocyst formation were analyzed by chi-square analysis, and data on mRNA expression were analyzed by one-way repeated-measures ANOVA. No differences in cleavage rates were observed between groups. There was no difference between the BSA group with or without s-HA in the percentages of embryos developed to the blastocyst stage at Days 7, 8, and 9 (38.3 and 38.1%, respectively). However, significantly (P < 0.05) less blastocysts developed in medium supplemented with BSA + FCS (18.3%) or with BSA + FCS + s-HA (27.4%). Synthetic HA added to the medium containing BSA significantly (P < 0.05) increased the level of expression of EGF-II and decreased (P < 0.05) the level of expression of BAX, SOX, and Ecad. On the other hand, presence of FCS significantly (P < 0.05) increased the level of SOX and decreased the level of IGF-II (P < 0.05), and the addition of s-HA to SOF containing FCS showed no effect on the level of transcription of any analyzed genes. The general fine structure of embryos cultured with s-HA regardless of protein supplement was conspicuously improved in comparison with the respective controls. It can be concluded that, within our culture system, addition of s-HA on Day 4 of culture to the SOFaa medium supplemented with BSA but not in combination with FCS showed a positive effect on embryo development and molecular composition of the embryos.

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Elimination of browning exudate and in vitro development of shoots in Pistacia vera L. cv. mateur and Pistacia atlantica Desf. Culture

Acta Societatis Botanicorum Poloniae ◽

10.5586/asbp.1999.004 ◽

2014 ◽

Vol 68 (1) ◽

pp. 21-24 ◽

Cited By ~ 3

Author(s):

M. I. Trujillo ◽

S. Mederos-Molina

Keyword(s):

Shoot Organogenesis ◽

Shoot Formation ◽

The Other ◽

Pistacia Vera ◽

In Vitro Development ◽

Pistacia Atlantica ◽

Murashige And Skoog Medium ◽

In Vitro Establishment ◽

Vitro Development

We report diminution and/or elimination of browning exudate followed by in vitro establishment of in Pistacia vera cv. mateur and Pistacia atlantica explants. Soaking P. vera cv. mateur explants prior to culture in L-cysteine HCl for 15 min (100 µM) inhibits blackening of the modified Murashige and Skoog medium - MS + 400 mg/l NH4NO3 - and of the explants; while shoot formation was increased. The browning in P. vera cv. mateur and P. atlantica explants dissolved when modified MS and Quoirin and Lepoivre - QL.4 - media were supplemented with activated charcoal (from 1 to 3 g l-1) and with 4 and 8 days of darkness. These treatments were enough to eliminate browning from the explants and to improve the shoots elongation, but symptoms of chlorosis were detected. On the other hand, AgNO3 (from 15 to 40 µ1V1) showed a very strong antibrowning effect on the medium and explants of P. atlantica. Thus shoot organogenesis was best achieved and the developing sturdy shoots had large and green leaves.

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