THE EFFECT OF OVARIECTOMY ON RABBIT BLASTOCYSTS

1962 ◽  
Vol 24 (2) ◽  
pp. 185-197 ◽  
Author(s):  
CECILIA LUTWAK-MANN ◽  
MARY F. HAY ◽  
C. E. ADAMS
Keyword(s):  
Mitotic Activity ◽  
Culture Media ◽  
Experimental Period ◽  
Carbonic Anhydrase Activity ◽  
Microscopical Examination ◽  
Fresh Weight ◽  
Stage Of Development ◽  
Embryonic Loss ◽  
Uterine Environment

SUMMARY The behaviour of rabbit blastocysts and their uterine environment after ovariectomy was studied; the experimental period after ovariectomy varied from 17 to 26 hr. Most of the experiments were concerned with blastocysts recovered from rabbits spayed at 6 days; a few relate to embryos from animals spayed on day 5 or 7 of gestation. Ovariectomy resulted in variable, often considerable, embryonic loss. Following bilateral ovariectomy at 6–6½ days, implantation on day 7 was completely prevented; those blastocysts which survived showed evidence of expansion and differentiation during the interval after ovariectomy. This provided an opportunity (1) to assess the stage of development, and mitotic activity, as well as certain biochemical characteristics, in embryos surviving ovariectomy, and (2) to use the free-lying 7-day-old blastocysts for culture experiments. It was found that mitotic activity went on in blastocysts after ovariectomy. Upon transfer to suitable culture media the unimplanted 7-day-old blastocysts continued to grow and differentiate, the most advanced stage observed at the end of a 24 hr. period of incubation in vitro being the development of blood islands, primitive groove, and head process. The fresh weight, and the content of bicarbonate and glucose of the 7-day-old unimplanted blastocysts differed little from those of normally implanted blastocysts of the same age; their content of lactate, however, was distinctly lower. Carbonic anhydrase activity after spaying was fully maintained in the endometrium which showed little involutional change either on gross or on microscopical examination.

In vitro analysis of glucose metabolism and embryonic growth in postimplantation rat embryos

Development ◽  
1987 ◽  
Vol 100 (3) ◽  
pp. 431-439 ◽  
Author(s):  
S.K. Ellington
Keyword(s):  
Glucose Metabolism ◽  
Embryonic Development ◽  
Glucose Uptake ◽  
Glucose Concentration ◽  
Culture Media ◽  
Rat Embryos ◽  
Embryonic Growth ◽  
Stage Of Development

The glucose metabolism and embryonic development of rat embryos during organogenesis was studied using embryo culture. Glucose uptake and embryonic growth and differentiation of 10.5-day explants (embryos + membranes) were limited by the decreasing glucose concentration, but not the increasing concentration of metabolites, in the culture media during the second 24 h of a 48 h culture. No such limitations were found on the embryonic development of 9.5-day explants during a 48 h culture although glucose uptake was slightly reduced at very low concentrations of glucose. From the head-fold stage to the 25-somite stage of development, glucose uptake was characteristic of the stage of development of the embryo and not the time it had been in culture. Embryonic growth of 9.5-day explants was similar to that previously observed in vivo. Glucose uptake by 9.5-day explants was dependent on the surface area of the yolk sac and was independent of the glucose concentration in the culture media (within the range of 9.4 to 2.5 mM). The proportion of glucose converted to lactate was 100% during the first 42h of culture then fell to about 50% during the final 6h. The protein contents of both the extraembryonic membranes and the embryo were dependent on the glucose uptake.

154 COMPARISON OF DIFFERENT CULTURE MEDIA AND INCUBATION METHODS ON CULTURING MURINE EMBRYOS IN VITRO USING STRAW AS A RECEPTACLE

2017 ◽  
Vol 29 (1) ◽  
pp. 185
Author(s):  
L. R. Madzhie ◽  
M. A. Raseona ◽  
L. P. Nethenzheni ◽  
O. Ajao ◽  
M. L. Mphaphathi ◽  
...  
Keyword(s):  
Developmental Stages ◽  
Culture Media ◽  
Uv Light ◽  
Fertilization Rate ◽  
Blastocyst Stage ◽  
Blastocyst Formation ◽  
Vitro Fertilization ◽  
Stage Of Development ◽  
Murine Embryos

In vitro fertilization in the straw system might increase the efficiency of fertilization and the quality of blastocyst formation as compared with micro-drops-IVF systems. The aim of the study was to in vitro fertilize mouse oocytes and culture the resulting zygotes in bi-gas incubator and in a goat vagina and compare the in vitro embryo developmental stages in TCM-199 and Ham’s F10 culture media until the blastocyst-stage of development. F1 generations (Balb C × C57) were used to harvest oocytes and spermatozoa. The fresh sperm were capacitated in different incubation methods (bi-gas incubator and in the vagina of a goat). A volume of 2–4 µL of Ham’s F10 containing capacitated sperm (~8 × 106 per mL) were placed into Ham’s F10 fertilization drops under the oil, containing 10 oocytes and penicillamine, hypotaurine, and epinephrine for enhancing sperm motility and penetration of oocytes. The same procedure was used with the TCM-199 medium and IVF drops without oil (both TCM-199 and Ham’s F10) for straw filling. The presumptive embryos in Ham’s F10 and TCM-199 were divided into different groups: first group were cultured in micro-drops, second group the embryos were aspirated in semen straws and placed in the incubator (incubator straws) for culture, and other straws were covered with a sponge and inserted in the vagina of a goat (vaginal straws) for culture. The resulted blastocysts were stained using Hoechst 33528 solution and blastomeres were counted on a fluorescent UV light inverted microscope at 400× magnification (Nikon Eclipse TI, Narishige Co., Ltd., Amityville, NY, USA). The results were analysed by 2 × 2 factorial designs and Student’s t-test was used to separate the mean. There was no statistical difference (P > 0.05) between the media and incubators on the stage of murine embryo development. The overall fertilization rate was 94 to 99%. The incubator straws with Ham’s F10 (80.5%) had the highest rate of embryos that reached the blastocyst stage, followed by incubator straws with TCM-199 (77.0%), and vaginal straws with Ham’s F10 (60.0%) had the lowest rate of embryos that reached the blastocyst stage. The overall mean number of blastomeres in the blastocyst stage of the embryos ranged from 85 ± 9 to 90 ± 9 cells in all receptacles and incubators. It was concluded that the fertilization and culturing of murine embryos are possible in straws incubated in a bi-gas incubator and in the goat vagina as an alternative method of fertilizing oocytes and culturing murine embryos. In addition, Ham’s F10 and TCM-199 can both be used to fertilize oocytes and culture murine embryos until blastocyst formation embryo in vitro, incubated in a bi-gas incubator or in the vagina.

scholarly journals THE ROLL OF SOME PLANT GROWTH REGULATORS ON SHOOTS MULTIPLICATION OF STEVIA PLANTS IN VITRO

2017 ◽  
Vol 48 (5) ◽  
Author(s):  
Al-Obaidy & Khierallah
Keyword(s):  
Plant Growth ◽  
Plant Growth Regulators ◽  
Growth Regulators ◽  
Culture Media ◽  
Stevia Rebaudiana ◽  
Shoot Multiplication ◽  
Dry Weight ◽  
Fresh Weight ◽  
Number Of Shoots

This research was conducted to study the effect of some plant growth regulators on in vitro shoots multiplication of stevia (Stevia rebaudiana Bertoni). The experiments included tests of various combinations of KIN with IBA or IAA in the shoot multiplication. Results indicated that KIN at 1.0 mg. L-1 plus 0.3 mg. L-1 of IBA produced the highest number of shoots (3.5 shoots) while KIN at 1.5 mg. L-1 plus IBA at 1.0 mg. L-1 produced the lowest shoot length (1.14 cm).  Hormone free medium produced the highest rate of the leaves number reached 28.56 leaves. KIN and IBA interaction increased fresh and dry weight significantly.   Treatment contained 2.0 mg -1 KIN plus 0.3 mg. L-1 IBA produced the highest fresh weight (1.739 g) while 0.5 mg. L-1 KIN and 0.3 mg. L-1 IBA produced the highest dry weight (0.822 g). As for the effect of interaction between the IAA and KIN it was significant in the number of shoots formed. Interaction between 1.0 mg. L-1 KIN with 0.1 mg. L-1IAA produced the highest number of shoots (3.8 shoots). Shoots length reached 8.10 cm in the media with 0.3 mg. L-1 IAA only. The highest fresh weight (1.267 g) was achieved with the interaction between 1.0 mg. L-1 KIN and 0.3 mg. L-1 IAA while 0.5 mg. L-1IAA without KIN produced the highest dry weight reached 0.138 g.  Shoots multiplication was improved by incorporation of the cytokinin TDZ in culture media. Shoots number, fresh and dry weights were increased significantly by adding 0.05 mg. L-1 of TDZ at present of 0.3 mg. L-1 of IBA giving 6.6 shoots, 0.974 g and 0.144 g respectively while shoots length decreased significantly as media without TDZ produced the highest shoots length reached 9.32 cm. The above results can adopt for the successful in vitro shoot multiplication of Stevia plants. 

Analyse biostatistique du développement cellulaire du tube pollinique, au cours de la gamétogenèse mâle expérimentale du Juniperus communis (Cupressacées)

1979 ◽  
Vol 57 (16) ◽  
pp. 1754-1760 ◽  
Author(s):  
E. Duhoux ◽  
P. D. M. Macdonald
Keyword(s):  
Culture Media ◽  
Pollen Grains ◽  
Quantitative Comparison ◽  
Juniperus Communis ◽  
Male Gametogenesis ◽  
Stage Of Development ◽  
Male Gametes ◽  
The Mean ◽  
Biostatistical Analysis

Male gametogenesis in gymnosperms proceeds through a series of well-defined stages, the time spent in each stage being variable. Pollen grains of Juniperus communis L. were allowed to develop in vitro and the numbers of pollen tubes in each stage of development were noted at subsequent times. A biostatistical analysis yields estimates of the mean durations of the stages, the rate of formation of the male gametes, and an upper bound on the rate of mortality, parameters which are virtually impossible to measure directly. The analysis allows a quantitative comparison of development rates in two different culture media.

98 EXPERIMENTAL TRANSFER OF BOVINE IVF-DERIVED 32-CELL STAGE EMBRYOS INTO THE UTERUS: ENVIRONMENTAL EFFECTS ON DEVELOPMENTAL CHARACTERISTICS

2016 ◽  
Vol 28 (2) ◽  
pp. 179
Author(s):  
M. Hoelker ◽  
D. Salilew-Wondim ◽  
F. Rings ◽  
D. Tesfaye ◽  
K. Schellander
Keyword(s):  
Culture Media ◽  
Uterine Cavity ◽  
Blastocyst Stage ◽  
Considerable Proportion ◽  
Bovine Embryo ◽  
Bovine Embryos ◽  
Cell Stage ◽  
Developmental Rates ◽  
Uterine Environment

Usually, in vitro-produced bovine embryos are cultured in vitro in static culture systems for 7 to 9 days in media composed according the oviducal fluid although it is well accepted that around Day 4.5–5 the bovine embryo enters the uterine cavity, providing environmental conditions different from the oviduct. Therefore, one has to raise the question whether changing culture media properties after Day 5 of culture could have beneficial effects on early development of bovine embryos. To answer that question, we transferred bovine IVF derived 32-cell stage embryos into the uterine cavity of synchronized recipients. All embryos had been matured and fertilized under routine standard conditions and were cultured in synthetic oviducal fluid supplemented with essential and nonessential amino acids (SOFaa) supplemented with either 0.3% fatty acid free bovine serum albumin (BSAfaf/Uterus) or 10% serum (serum/uterus) at 38.5°C, 5% O2, and 5% CO2 in humidified air prior transfer into the uterine environment, allowing further development to the blastocyst stage within the physiological environment prior recollection at Day 7 by routine uterine flushing followed by comparison with statically in vitro-developed embryos cultured in media supplemented with serum (serum/serum group) or BSAfaf (BSAfaf/BSAfaf group). All in all, a total of 1031 in vitro-derived 32-cell stage embryos were transferred to 21 synchronized Simmental recipient heifers. Of these, a total of 680 embryos (66%) could be recollected at Day 7. Embryos of the serum/serum group reached a higher blastocyst rate compared with embryos of the BSAfaf/BSAfaf group (68% v. 41%; P < 0.05, ANOVA, Tukey test), whereas the developmental rate to the blastocyst stage did not differ after 9 days of in vitro culture, indicating higher developmental kinetics of bovine 32-cell stage embryos when culture media is supplemented with serum. Moreover, embryos of the serum/uterus group reached significantly lower developmental rates to the blastocyst stage until Day 7 compared with embryos of the serum/serum group (12.9% v. 68.4%). Likewise, embryos in the BSAfaf/uterus group reached significantly lower developmental rates to the blastocyst stage until Day 7 compared with embryos in the BSAfaf/BSAfaf group (16.0% v. 40.1%). When allowed to develop for additional 48h in vitro, developmental rates to the blastocyst stage at Day 9 were still higher in BSAfaf/BSAfaf treatment compared with the BSAfaf/uterus treatment (91.4% v. 74.4%) and the serum/serum treatment compared with the serum/uterus treatment (92.5% v. 56.0%). Taken together, the results of our study demonstrate that uterine transfer of bovine 32-cell stage embryos results in reduction of developmental kinetics as well as lower developmental rates compared with embryos statically cultured in vitro. That might indicate, that a considerable proportion of bovine 32-cell stage embryos might not be able to adapt to the uterine environment.

scholarly journals The influence of Cinnamomum zeylanicum in plant tissue culture experiments of Gardenia jasmenoides Ellis

2013 ◽  
Vol 10 (4) ◽  
pp. 1102-1107
Author(s):  
Baghdad Science Journal
Keyword(s):  
Root Formation ◽  
Culture Media ◽  
Shoot Proliferation ◽  
Naphthalene Acetic Acid ◽  
Alcoholic Extract ◽  
Fresh Weight ◽  
Benzyl Adenine ◽  
Cinnamomum Zeylanicum ◽  
Positive Effect

Cinnamon plant is considered one of important medicinal plants because it is rich with many active compounds. This research is aimed to study possible effects of extract in culture media of Gardenia jasmenoides. Alcoholic extract was prepared from the bark of cinnamon at different concentrations (0.0, 1.0, 2.0) mg/L, then added to culture media to notice the effect of these concentrations on the growth and development of tissues and organs of Gardenia jasmenoides Ellis in vitro. Results showed the positive effect of increasing callus fresh weight and shoot proliferation from single nodes with presence of plant regulators, 5.0 mg/L Naphthalene acetic acid (NAA) and 3.0 mg/L Benzyl adenine (BA). Results showed that extract has a slight effect on root formation with the presence of plant regulators or when it is alone.

scholarly journals A Comparison between Synthetic and Organic Iron Chelators on in Vitro Growth of Nicotiana tabacum Callus Cultures

HortScience ◽  
1996 ◽  
Vol 31 (4) ◽  
pp. 630g-631
Author(s):  
Lisa C. Berg ◽  
Henry R. Owen
Keyword(s):  
Nicotiana Tabacum ◽  
Culture Media ◽  
Basal Medium ◽  
Callus Cultures ◽  
Callus Growth ◽  
Fresh Weight ◽  
Iron Chelate ◽  
Tissue Culture Media ◽  
The Media

Nicotiana tabacum callus growth (fresh weight) was measured after culture in the light (16-hour photoperiod) or in darkness for four different culture media, differing in iron chelate type or concentration. All media contained MS basal medium supplemented with 30 g·L–1 sucrose, 2 mg·L–1 IAA, 0.2 mg·L–1 KIN, and 7 g·L–1 agar, pH 5.8. Three of the media contained iron-metalosate (Albion Laboratories), an organic iron chelate, at 100, 200, and 400 micromolar concentrations, and the fourth medium contained 100 μm Fe-EDTA. Twenty-five culture tubes were prepared for each of the 4 different media concentrations and 2 light treatments (8 treatments total). A 1-cm3 callus explant was used for each treatment and cultured for 56 days at 20°C. About 20-fold increases in callus fresh weight were observed for cultures incubated in light or in darkness. In addition, callus growth was not significantly affected by iron chelate type, suggesting the potential utility of this organic chelator in tissue culture media to alleviate potential problems of light-induced EDTA instability and subsequent IAA inactivation. These cultures are being maintained to examine the influence of iron chelate type on organogenesis.

scholarly journals Initial segmentation patterns of microspores and pollen viability in soybean cultured anthers: indication of chromosome doubling

2004 ◽  
Vol 47 (5) ◽  
pp. 703-712 ◽  
Author(s):  
Milena Barcelos Cardoso ◽  
Eliane Kaltchuk-Santos ◽  
Elsa Cristina de Mundstock ◽  
Maria Helena Bodanese-Zanettini
Keyword(s):  
Chromosome Doubling ◽  
Culture Media ◽  
Pollen Viability ◽  
Pollen Grains ◽  
Chromosome Counts ◽  
Viable Pollen ◽  
Stage Of Development ◽  
Initial Segmentation ◽  
Bud Size

Anthers obtained from flowers buds of soybean cultivar IAS-5 were cultured in two basal culture media (B5 and B5 long). Cytological examinations of the in vitro anthers were performed during the first 20 days of culture to assay the viability (by propionic-carmine and fluorescein diacetate tests) and the stage of development of pollen grains. The frequencies of viable pollen grains varied significantly between bud sizes on the propionic-carmine analysis. The basal culture media and bud size had no clear effect on the frequencies of binucleate symmetrical and multinucleate pollen grains. Chromosome counts of metaphasic microspores throughout the culture period showed microspores with higher ploidy level in addition to normal chromosome number (n=20).

168 CORRELATION OF DEVELOPMENT KINETICS AND SEX OF IN VITRO-PRODUCED BOVINE EMBRYOS

2012 ◽  
Vol 24 (1) ◽  
pp. 196
Author(s):  
A. R. Buzzo ◽  
A. R. Pupulim ◽  
J. Mazucheli ◽  
F. V. Meirelles ◽  
I. P. Emanuelli
Keyword(s):  
In Vitro Culture ◽  
Culture Medium ◽  
Culture Media ◽  
Proteinase K ◽  
Bovine Embryos ◽  
Male And Female ◽  
Stage Of Development ◽  
Males And Females ◽  
Kinetics Of

Approaches to improve the culture medium for in vitro production (IVP) of bovine embryos have been continuous because of the high commercial demand and a portion of this attempts the production of female cattle (dairy cows and stud cattle). However, in some embryonic in vitro culture systems, the development kinetics is faster in male than in female embryos (Avery 1992 Mol. Reprod. Dev. 32, 265–70; Xu 1992 Mol. Reprod. Dev. 31, 249–50). The aim of this work was to relate the kinetics of blastocyst expansion with the production rates of male and female embryos. Cumulus–oocyte complexes (n = 917; classes I and II) of cows from a slaughterhouse were matured with TCM-199 bicarbonate and 10% FCS (38.5°C, 5% CO2) for 24 h and fertilized with frozen-thawed semen in TALP-IVF medium for 18 h. Presumptive zygotes were culture in SOF medium supplemented with 10% FSB (5% O2, 38.5°C). Seven days after IVF, embryos were divided in 2 groups according to their kinetic stage of development: nonexpanded blastocysts (n = 175), or hatched and expanded blastocysts (n = 146). Hence, embryos were individually frozen in LN and stored in cryotubes. After thawing, Proteinase K (16 mg mL–1) was added to each tube and the tubes were incubated for 60 min at 37°C. Proteinase was denatured at 98°C for 10 min and the contents of each tube were divided into 2 samples (A and B) and subjected to the PCR technique. Two pairs of primers for the specific sequence of the Y chromosome were used to amplify the sequence of 210 and 250 bp for the male bovine and 1 pair of primers was used for the autosomal bovine sequence with a 280-bp fragment. Female embryos with a 280-bp product were observed in sample A and none were observed in sample B. The presence of 2 amplicons (280 and 210 bp) in sample A and 1 amplicon of 250 bp in sample B indicated that the embryo was male. A chi-square test was used to evaluate homogeneity. An analysis of the percentage of males and females between the experimental groups was performed by logistic regression and significance was considered when P < 0.05. There was no difference in the proportions of males and females in the nonexpanded blastocyst group (49.71 and 50.29%; P > 0.05). In the hatched and expanded blastocyst group, the proportion of males (65.75%) was statistically different from the proportion of females (34.25%); that is, the chance of the embryo being male was twice as high (P < 0.0038). These results suggest that there is a difference in the kinetics of embryo development between male and female embryos and that blastocyst expansion can point that out. In vitro culture media with FCS support the development of expanded male blastocysts. Further research in culture medium modifications (FCS, the energy source, amino acids and others) are needed to respond to the trend in the production of sex-defined embryos.

A modified method for assessment of the morphological stage of development as a predictor of alfalfa herbage chemical composition and nutritive value

2013 ◽  
Vol 151 (4) ◽  
pp. 590-598 ◽  
Author(s):  
A. BOŽIČKOVIĆ ◽  
G. GRUBIĆ ◽  
J. VERBIČ ◽  
T. ŽNIDARŠIČ ◽  
N. DJORDJEVIĆ ◽  
...  
Keyword(s):  
Chemical Composition ◽  
Nutritive Value ◽  
Development Stage ◽  
Individual Growth ◽  
Coefficient Of Determination ◽  
Fresh Weight ◽  
Growth Cycles ◽  
Stage Of Development ◽  
Average Development

SUMMARYThe aim of the current work was to investigate the possibility of modifying the existing mean stage by weight (MSW) system for evaluating the average development stage in alfalfa. The modification was performed with the aim of providing a simplified system that may be used to evaluate the alfalfa development stage and to predict its nutritive value for ruminants. The suggested modification consists of designating an MSW value on the basis of the fresh weight of all morphological stages in a fresh green plant, as opposed to the original method which is based on weighing all morphological stages dried at 65 °C. The investigation was done on 141 samples of one alfalfa cultivar, collected from the same location during the first three growth cycles: spring growth, the first and the second regrowth. On all collected samples the following characteristics were determined: MSW, modified MSW (mean stage by fresh weight (MSFW)), crude protein (CP), neutral detergent fibre (NDF), acid detergent fibre (ADF), acid detergent lignin (ADL), crude ash (CA) and in vitro organic matter digestibility (IVOMD). For these characteristics of chemical composition (apart from CA) and nutritive value the regressions were calculated for their prediction based on MSW and MSFW. The regressions were derived for individual growth cycles and all cycles combined. A trend for an increase in the coefficient of determination (R2) was identified as well as a decrease in root-mean-square error (RMSE) for all equations derived for all investigated characteristics from the spring growth to the second regrowth. A deviation from this trend was observed only in equations derived for IVOMD. A very high correlation was observed between MSW and MSFW (r=0·999). The determined R2 and RMSE were very similar within the same growth cycle in all regressions for prediction of chemical composition and nutritive value derived for MSW and MSFW. Based on the results of this investigation the MSFW appears to be a quick and accurate method for determining the average development stage in alfalfa which can therefore be recommended for both scientific research and practical field use, as well as for prediction of its chemical composition and nutritive value.

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