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At least 1,423 records · Page 79Linked to original sources
PubMed2006

Survival of mouse embryos after vitrification depending on the cooling rate of the cryoprotectant solution.

The aim of the study was to determine the relationship between the rate of cooling of eight-cell mouse embryos to the temperature of liquid nitrogen (-196 degrees C) and their developmental capacity after thawing on the basis of their ability to leave the zona pellucida ('hatching') during in vitro culturing. Eight-cell embryos were obtained from superovulated female mice and divided into three experimental and one control group. Embryos from the experimental groups were cryopreserved by the vitrification method using ethylene glycol as cryoprotectant. The vitrification protocols used in the study differed in the rate of cooling of the cryoprotectant solution. Embryos from the first group were frozen in conventional 0.25-ml plastic straws, those from the second group in pipetting 'tips', and embryos from the third group, placed in vitrification solution, were introduced dropwise directly into liquid nitrogen. The control group of embryos was cultured in vitro without freezing in a culturing medium in an environment consisting of 95% air and 5% CO2. The developmental capacity of thawed embryos was assessed on the basis of their ability to leave the zona pellucida ('hatching') after three days of in vitro culturing. In the control group 95.1% of embryos 'hatched'. A significantly higher number of embryos that 'hatched' after thawing was observed in the group introduced dropwise directly into liquid nitrogen (60.0%) compared to the group frozen in pipetting 'tips' (37.9%). The group frozen in straws yielded significantly the lowest proportion of 'hatching' embryos (8.1%). These results showed that increasing cooling rates during vitrification of embryos improved their survival.

Animals↗
PubMed1994

Effects of blastocoelic expansion and plasminogen activator activity on hatching and zona pellucida solubility in bovine embryos in vitro.

Two experiments were conducted to evaluate factors affecting zona pellucida (ZP) solubility in bovine embryos during hatching in vitro. In Exp. 1, the relationship between blastocoelic expansion and ZP solubility was determined. Day-6 embryos (n = 42) with good or excellent quality grades were cultured (Cultured embryos), whereas embryos of fair quality (Not-cultured embryos) were immediately placed in .2% SDS, and the time required for complete dissolution of the ZP (ZPDT) was determined. For Cultured embryos, ZPDT was determined after 192 h of in vitro development. Zona pellucida dissolution time was greater (P < .05) in Not-cultured embryos than in Cultured embryos and negatively correlated (P < .01) with changes in embryonic surface area. In Exp. 2, the effects of suppressing plasminogen activator (PA) activity and blastocoelic expansion on hatching and ZP solubility were examined. Day-6 embryos (n = 99) were cultured in medium containing 0, 10, or 100 IU/mL of human PA inhibitor-2 (PAI-2) and incubated for 24 to 44 h in medium containing 0, .1, or .5 nM ouabain. Percentages of embryos hatching were not different (P > .05) among 0, 10, and 100 IU/mL of PAI-2; however, more (P < .05) embryos hatched after exposure to 0 and .1 nM ouabain than to .5 nM ouabain. Embryonic PA activity was suppressed (P < .05) by PAI-2, whereas exposure to ouabain did not affect (P > .05) PA activity. Mean ZPDT did not differ (P > .05) following culture in medium containing PAI-2 or exposure to ouabain.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1997

Evidence that injection of vitamin A before mating may improve embryo survival in gilts fed normal or high-energy diets.

The hypothesis was that administration of vitamin A before ovulation would improve embryo survival in gilts fed a high-energy diet intentionally to reduce embryo survival. Forty crossbred ([Landrace x Large White] x [Duroc x Hampshire]) gilts were fed control (5.5 Mcal ME/d) or high-energy (11.0 Mcal ME/d) diets from 7 d after second estrus until 11 to 12 d after third estrus. Gilts in each dietary group received (i.m.) corn oil or retinyl palmitate (1 x 10(6) IU, vitamin A) on d 15 after second estrus and were mated at third estrus. Blood for determination of progesterone and estradiol was collected twice daily. The uterus and ovaries were removed on d 11 or 12 after third estrus for assessment of number of CL, and number, size and aromatase activity of embryos. Neither diet nor vitamin treatment affected number of CL. The high-energy diet exerted a negative effect on number of embryos (P = .09) and embryo survival (P = .07), whereas vitamin A exerted a positive effect on number of embryos (P = .07) and embryo survival (P = .08). The high-energy diet increased variation in embryo diameter, whereas vitamin A reduced variation in diameter and increased average diameter. Neither diet nor vitamin treatment influenced aromatase activity of embryos. Diet and vitamin treatment interacted with day to influence serum progesterone, but not estradiol. Injecting vitamin A before estrus restored embryo survival to normal levels in gilts fed high-energy diets, and this may be attributable to decreased variation in size of embryos.

Animals↗
PubMed2003

Evaluation of the uterine environment and embryos of prepubertal gilts.

A series of three experiments was conducted to test the functional status of the uterus and embryos in prepubertal gilts. In Exp. 1, gilts were induced to ovulate by treating with gonadotropins followed by hCG 72 or 96 h later, and were artificially inseminated 24 h after hCG. Five of the 10 gilts treated at 120 d of age, but none of the gilts treated at 100 of age, maintained pregnancies. We next tested the function of the uterine environment by transferring embryos from postpubertal females into gilts of various ages that had been induced to ovulate but not inseminated (Exp. 2). Pregnancy rate at d 50 of gestation was 44% (4/9) for 100-d-old recipients, 67% (2/3) for 140-d-old recipients, and 60% (3/5) for postpubertal recipients (P > 0.20). Therefore, uteri of 100-d-old gilts are able to maintain pregnancies with conceptuses from postpubertal gilts. In Exp. 3, embryos from 100-d-old and postpubertal gilts were transferred into postpubertal recipients. Uterine horns of recipients were surgically separated before transfer, and embryos from 100-d-old and post-pubertal females were transferred to opposite horns of some recipients (experimental). Other recipients received embryos from postpubertal females in both uterine horns (control). When examined on d 50 to 60 of gestation, three of five control gilts were pregnant and three of seven experimental gilts were pregnant (P > 0.50). In experimental recipients, the survival of embryos from 100-d-old gilts was 38% (8/21) compared to 57% (15/26) for embryos from postpubertal gilts (P > 0.30). Because all uterine horns of pregnant recipients contained fetuses, these results support the hypothesis that embryos from 100-d-old gilts are able to initiate and maintain pregnancies in the uteri of postpubertal gilts. Therefore, the uterine environment of 100-d-old gilts provides an environment that supports development of embryos produced by postpubertal gilts, and the embryos produced by 100-d-old gilts can survive and develop in the uteri of postpubertal gilts. It was only the combination of embryos and uteri of 100-d-old gilts that did not permit pregnancy to be maintained.

Age Factors↗
PubMed1985

Effect of slow cooling end point temperature on survival of frozen bovine embryos.

One hundred sixty-two embryos were collected from superovulated crossbred beef cattle 7 to 8 d after the onset of estrus. Embryos were frozen in modified Dulbecco's phosphate-buffered saline supplemented with 20% heat-inactivated fetal calf serum (PBS + FCS) and dimethylsulfoxide (DMSO), which was added in three steps to a final concentration of 1.5 M. Embryos were placed in .25 ml of 1.5 M DMSO in PBS + FCS in 1-ml glass ampules and cooled at 1.0 C/min from ambient temperature to -7 C, seeded and then cooled at .3 C/min to -19, -26, -33, -38, -43, -50 or -57 C before immersion (plunging) in liquid nitrogen. Ampules were thawed in 25 C water, and DMSO was removed in six steps at .25 M increments. 10 min/step. After removal of DMSO, embryos were cultured 24 h in PBS + FCS and then fixed and stained. Just after thawing, embryos for which slow cooling was terminated at -50 C were of lower (P less than .05) morphological quality than other groups. After removal of cryoprotectant, embryos from both the -19 and -50 C treatments had deteriorated more (P less than .05) than had embryos from other treatments. After 24-h culture, embryos slow-cooled to -19, -26 and -50 C had a lower rate of survival (P less than .05) than did embryos from -33, -38, -43 and -57 C temperatures. Embryos slow-cooled to -33, -38 and -43 C showed a higher percentage of healthy nuclei than did embryos slow-cooled to -19, -26 and -50 C.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1985

Bovine embryo transfer pregnancies. I. Abortion rates and characteristics of calves.

Data were obtained from 1,908 pregnancies resulting from bovine embryo transfer procedures. Responses examined included sex ratio, fetal, neonatal and preweaning death losses, birth weight and calving assistance. The sex ratio for 1,751 embryo transfer calves examined was 51.11% males. Cows older than 10 yr that had become repeat breeders produced more (P less than .05) male calves than other donors. Breed of embryo, age and quality of embryos at the time of transfer, embryo storage time from collection to transfer, asynchrony of recipient with donor estrus and number of palpable corpora lutea in superovulated donors were not related to sex ratio (P greater than .05). The abortion rate between 2 and 3 mo of gestation in embryo transfer recipients was 3.15%, and between 3 to 7 mo, 2.14%. Neonatal and preweaning losses for 1,682 calves with complete information were 1) congenital defects, .54%; 2) death due to premature birth (7 to 8 mo of gestation), .18%; 3) dystocia-related deaths, 2.38%; 4) deaths of unknown causes at birth, 2.14%; 5) deaths of unknown causes from 24 h after birth to weaning, 1.43%; 6) deaths due to calfhood diseases, 1.25% and 7) deaths due to environmental factors, 1.13%. Total losses of 2-mo pregnancies due to abortion or death of calves or recipients were 14%. Birth weight of embryo transfer calves changed .29 kg/d of deviation from average gestation length (P less than .005) for pregnancies within breeds. Birth weight was also affected (P less than .005) by donor breed and recipient breed and age. Male calves averaged 2.19 kg heavier (P less than .005) than females. Calving assistance was affected by donor breed; Angus calves required the least assistance (P less than .005). Hereford, Holstein and Limousin calves were similar and intermediate; Simmental calves needed the most calving assistance. Recipient breed and age influenced calving ease, with younger recipients of Angus and Hereford descent requiring more assistance (average calving score, 2.1) than both cow (1.3) and heifer (1.5) recipients of the larger Continental European breeds. Characteristics of 305 non-embryo transfer calves were not significantly different from 185 embryo transfer calves from the same farms. We conclude that embryo transfer calves did not differ from the non-embryo transfer population in any of the characteristics studied.

Abortion, Veterinary↗
PubMed1982

Long-term shell-less culture of turkey embryos.

A technique is described for the long-term shell-less culture of turkey embryos. Cultured embryos demonstrated a similar rate of growth through 17 days of total incubation (4 days in ovo plus 13 days in culture) compared to embryos incubated in ovo. Thereafter, the cultured embryos grew at a reduced rate compared to their in ovo counterparts. Cultured embryos also exhibited elevated hematocrit values but reduced serum protein levels compared to the embryos incubated in ovo. Serum zinc, copper, and iron concentrations were similar for both groups of embryos, although each element reached a maximum concentration at a different point during incubation. Embryos incubated in ovo demonstrated a sustained elevation of serum calcium concentration from day 16 of incubation that was absent in the cultured embryos. The significance of eggshell calcium stores to embryo growth is discussed. Shell-less avian embryo culture represents a potentially useful technique for the study of trace element metabolism during avian embryonic growth and development.

Animals↗
PubMed1999

Comparison of the expression of specific cell surface epitopes on in vitro fertilized and parthenogenetic bovine embryos.

Parthenogenetic embryos, which are produced by the spontaneous or artificial stimulation of the oocyte, partially develop in the complete absence of the male gamete but fail to produce live young in many mammalian species. The identification of developmentally regulated molecules on the cell surface of embryos has implicated their possible role in cell interactions during embryogenesis and differentiation. In this study the expression patterns of four stage-specific cell surface antigenic determinants (TEC-1, -2, -3, and -4) were investigated in both parthenogenetic and in vitro fertilized bovine embryos. When compared to embryos produced using in vitro fertilization methods the parthenogenotes, although appearing morphologically normal, differed markedly in their TEC epitope pattern of presentation. TEC-1, -2, -3, and -4 epitope presentation on in vitro fertilized embryos occurred during specific stages of preimplantation development. TEC-1 and -2 presentation was detected on oocytes and blastocysts only, TEC-3 on morulae and blastocysts, and TEC-4 on oocytes through to 8-cell embryos, with all subsequent stages negative. Parthenogenetic embryos did not show TEC-1, -2, or -3 epitope presentation whereas the TEC-4 epitope was present throughout the developmental period examined. Enzymatic cleavage of sialic acid residues on in vitro fertilized and parthenogenetic embryos resulted in presentation of the TEC epitopes during all the embryonic stages. Western blot analysis of the embryos showed the TEC epitopes to be present on all the embryonic stages examined. This study suggests the mechanisms responsible for control and presentation of each of the TEC epitopes may not be functioning the same in parthenogenetic embryos that undergo changed glycosylation or deglycosylation resulting in altered patterns of sialylation. The study also shows TEC epitope presentation may prove to be a useful indicator of parthenogenetically activated bovine embryos. J. Exp. Zool. 284:392-400, 1999.

Animals↗
PubMed1999

[Diminishing the risk of multiple pregnancies in in vitro fertilization: from selective transfer of two embryos to that of one blastocyst?].

The risk of multiple pregnancy after IVF needs to be drastically reduced. Several policies can be applied including the transfer of a maximum of three embryos to all patients, the fertilization of a maximum of three oocytes or a selective reduction of the number of transferred embryos. The first policy previously applied at the Fertility Clinic at Erasme Hospital until 1996, transferred two good quality embryos to patients with at least three good embryos. If this policy demonstrated that patients with two transferred embryos had similar chances of pregnancies compared to patients with three transferred embryos, it failed to sufficiently decrease the number of multiple pregnancies. The second policy applied since 1997, transferring a maximum of two average or good embryos to all patients aged under 35 years and with less than 3 previous attempts, demonstrated that while preserving the chances of pregnancy for these patients, it decreased by 20% the number of multiple pregnancies and almost eliminated triplets. With the improvement of culture media, it is now possible to culture embryos in vitro for a longer period and therefore transfer embryos with proven viability at a time corresponding more to in vivo physiological conditions. The implantation rates for these embryos, for patients with at least 4 previous attempts can reach 40%. If these results persist, it would be possible to transfer blastocysts to all patients and perhaps move on to the replacement of a single embryo, a policy that will practically eradicate all multiple pregnancies.

Adult↗
PubMed2000

Lack of physiological plasticity in the early chicken embryo exposed to acute hypoxia.

By exposing chicken embryos to hypoxia (10%) acutely (2, 4, and 6 hr) during early development (2, 3, and 4 days) we tested the hypothesis that hypoxia has an impact on embryonic growth and impairs cardiac development at the time cardiac morphogenesis is taking place. After the hypoxic perturbation, the embryos were allowed to develop until day 9, when embryo mass, heart mass, and rate of oxygen consumption were recorded. Four-day-old embryos exposed to 6 hr of hypoxia showed an increased mortality (38.9% versus 18% for controls), indicating the immediate effect of hypoxia on survivability. While only 8% of the controls displayed morphological abnormalities, 3- and 4-day-old embryos exposed for 6 hr showed more frequent developmental abnormalities (25% and 30% respectively). No significant differences in embryo or heart mass were found except in 4-day-old embryos exposed for 2 hr. Mass-specific oxygen consumption was not different between controls and embryos exposed to hypoxia at 2 or 3 days of development, but it was increased in 4-day-old embryos exposed for 4 hr (P < 0.05). These results suggest that an acute hypoxic episode does not have an impact when occurring very early in development (days 2 or 3). However, when the hypoxic episode occurs on day 4, survivability is largely decreased. Considering the lack of permanent effects on the surviving embryos, we suggest that the early embryo resorts to a simple strategy of death or survival, and the individual capacity for survival must be based on interindividual differences rather than the existence of compensatory mechanisms. J. Exp. Zool. 286:450-456, 2000.

Animals↗
PubMed2002

Expression of microinjected reporter gene lacZ during first cleavages of rabbit embryos.

The growing use of reporter genes in a model transgenic system has been a fundamental approach of biology, but the strategy of transgenic embryo selection prior to transfer to foster mothers may greately increase the efficiency of transgenic livestock production. This study was conducted to assess the possibility of beta-galactosidase (beta-gal)-labeled transgenic rabbit embryo production. Rabbit zygotes were obtained from superovulated females after mating. Zygotes were microinjected into male pronuclei with pCMV-lacZ or SV40-lacZ constructs; while some embryos were co-injected with the scaffold attachment sequences--SAR. Embryos from control non-injected and microinjected groups were cultured in vitro. After 24, 48, 72, or 96 h of culture the embryos were stained with X-gal for beta-galactosidase. Transgenic embryos produced by pronuclear injection showed a discrete pattern of beta-galactosidase expression. The percentage of transgenesis with pCMV-lacZalone was 1.5, but with SAR sequences it increased to 4.2. In the case of SV40-lacZ construct, the efficiency of transgenesis was 2.3% and 4.1%, respectively. The mosaicism was 66.7% for all embryos injected with both constructs with or without SAR. The highest numbers of 100%-transgenic (non-mosaic) embryos were found in the group co-injected with SV40-lacZ and SAR. Transgenesis was seen as early as 24 h after injection, in four-cell embryos. Most of the microinjected embryos showed delayed development as compared with control. It was concluded that lacZ may serve as a reliable reporter for early transgenic embryo selection in order to produce transgenic animals.

Animals↗
PubMed1992

Selective assisted hatching of human embryos.

Overall results of assisted hatching by zona drilling using acidic Tyrode's solution performed during three randomised trials in 330 in vitro fertilisation (IVF) patients are presented. It was demonstrated retrospectively and prospectively that assisted hatching by zona drilling was effective in embryos with thick zonae (> 15 microns). This procedure is called selective assisted hatching. In order to investigate whether the success rate of embryos with thin zonae (< 13 microns) can be improved further, a fourth trial was executed in 40 consenting patients. Embryos with thin zonae were left intact in one group (control), while the outside of zonae of similar embryos were thinned with acidic Tyrode's solution. Results thus far indicate that embryos with thin zonae do not benefit from this technique. Embryonic implantation (fetal heartbeat per embryo) was high (26%-27%) in both arms of the trial, probably as a result of selective zona drilling of low prognosis embryos with thick zonae. A method is presented for quantifying zona hardening in human embryos. The exposure to acidic Tyrode's solution of each embryo was expressed as a function of the duration to pierce the zona and the diameter of the needle. Preliminary findings suggested that embryonic viability is correlated with zona hardening. In order to test the hypothesis that extracellular fragments may affect embryonic viability, small amounts of fragments were removed from embryos during assisted hatching. The pregnancy rate in 36 patients with extracted fragments was relatively high (41%) considering the poor morphology of the embryos involved.

Adult↗
PubMed1991

Vitrification of intact and half 8-cell mouse embryos injected with DNA at 2-cell stage.

Two-cell embryos were collected from (C57BL/6 x CBA)F1 mice, microinjected with foreign DNA and transferred into temporary recipients. Intact and half 8-cell embryos recovered from oviducts 24 h later were frozen in microdrops of vitrification medium and stored in liquid nitrogen for 2 to 30 days. Development of frozen/thawed embryos was assessed by in vitro culture or transfer into day-1 recipients. From 1200 2-cell embryos 77% intact and 20% half embryos were obtained one hour after DNA injections. After transfer of 600 intact and 300 half embryos into temporary recipients, 77% intact and 73% half 8-cell embryos were recorded, respectively. Survival and in vitro development of frozen/thawed DNA-injected embryos were high, as 97% intact and 91% half 8-cell embryos developed to morulae or blastocysts. After transfer of 300 intact and 150 half 8-cell embryos into day-1 recipients, 99 (33%) and 19 (13%) young were born. None of the mice born from DNA-injected and cryopreserved embryos integrated injected genes.

Animals↗
PubMed1987

Calcium regulation in the embryonic chick. III. Calcium and phosphate in serum and allantoic fluid of normal and shell-less embryos.

Calcium metabolism of chicken embryos was profoundly affected by incubation in shell-less culture, but phosphate metabolism was largely undisturbed. Shell-less embryos exhibited hypocalcemia and hypocalciuria relative to normal embryos but had similar levels of phosphate in serum and allantoic fluid. The concentration of calcium in allantoic fluid declined during incubation in both groups, owing largely to accompanying increases in allantoic volume, but total amounts of calcium in the allantois did not vary with time. Both normal and shell-less embryos maintained higher concentrations of calcium in serum than in allantoic fluid, with shell-less embryos maintaining a larger gradient between serum and allantoic compartments. In contrast, serum and allantoic concentrations of inorganic phosphate increased over time in both normal and shell-less embryos, and both groups maintained generally higher concentrations of inorganic phosphate in the allantoic sac than in serum. Treatment of embryos with parathyroid hormone had no effect on calcium and phosphate metabolism. Embryos maintained in shell-less culture grew more slowly than those incubated normally and consequently had a dry mass about half that of normal embryos on day 18. Shell-less embryos also exhibited abnormalities in fluid balance, which were reflected in their inability to maintain normal allantoic volume and in their higher relative hydration compared to embryos incubated in ovo.

Allantois↗
PubMed1985

Manipulation of gametes and embryos in the pig.

Several manipulation techniques including nuclear injections, nuclear transplantation, embryo splitting, chimaera production and sperm injection are discussed with special reference to their application in the pig. The nuclear injection technique is likely to be of greatest use for gene transfer. Gene transfer is feasible for pig embryos, but it is very inefficient. Efficiencies of various steps in the successful production of transgenic offspring as summarized from several published references are as follows: immediate survival of embryos following injection (60%), offspring produced from injected embryos (15%), offspring with an integrated foreign gene (24%) and offspring expressing the integrated foreign gene (60%). The overall efficiency is about 1%. Potential uses for nuclear transplantation are for the production of gynogenetic and androgenetic embryos and for cloning embryos. The feasibility of producing androgenetic and gynogenetic embryos will depend on whether in the pig, as in the mouse, the paternal and maternal genomes function differently. The feasibility of cloning by nuclear transplantation will depend upon the timing of developmental events in pig embryos and the development of methods for reprogramming nuclei. Pig embryos have been cloned by embryo splitting. Chimaera production and fertilization by sperm injection are likely to be feasible for pig embryos but have not yet found application.

Animals↗
PubMed1985

Effect of migration, distribution and spacing of pig embryos on pregnancy and fetal survival.

Pig embryos enter the uterus from the oviduct about 48 h after ovulation at about the 4-cell stage. They remain near the tip of the uterine horn until about Day 6 when they migrate towards the body of the uterus. By Day 9 some embryos have entered the horn opposite the one of origin and continue migrating until Day 12. At Day 12 embryos can no longer successfully move to a different site. The critical signal for recognition of pregnancy occurs at Day 12 at the same time that embryos are elongating at a rapid rate. The distribution of embryos throughout the length of the horns has occurred by this time and spacing accompanied by uterine growth takes place. The speed of migration is not influenced by the number of embryos but may be affected by secretions from the embryos acting on the uterus. A significant section of unoccupied uterus at Day 12 will prevent continuance of pregnancy regardless of the number of embryos present in the occupied section. When the number of embryos is so few as to not occupy the uterus fully, pregnancy will not continue. Removal of embryos from a significant section of the uterus after Day 14 does not stop an existing pregnancy. Embryos are spaced nearly equidistant from each other with the absolute space between fetuses dependent on the total uterine space available. The position of the fetus in the uterus affects the spacing between fetuses; fetuses at the tip of the horn before Day 25 have the greatest space with the space decreasing from the tip to the body.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Viability of fresh and frozen-thawed biopsied bovine embryos.

Bovine embryos were biopsied using a simplified splitting technique and frozen-thawed according to a standard method with glycerol as cryoprotectant. The viability of fresh and frozen-thawed biopsied and intact embryos were evaluated after in vitro culture, by means of fluorescence test or following transfer to recipients. The survival rates after in vitro culture of fresh intact and biopsied embryos and of frozen-thawed intact and zona free embryos were not significantly different (70%, 60%, 68% and 52%, respectively), but significantly reduced for biopsied frozen-thawed embryos (16%) (p < or = 0.05). The pregnancy results after transfer of biopsied frozen-thawed embryos were also significantly lower (8%) compared to fresh biopsied embryos (39%) (p < or = 0.05). Both intact and biopsied embryos fluoresced after incubation with diacetylfluorescin but with higher intensity for the intact embryos. It is suggested that the reduced survivability for the frozen-thawed biopsied embryos might be caused by combined effects of the loss of the zona pellucida and the reduction of cells as a result of the simplified biopsy technique. It is concluded that improved biopsy and/or freezing techniques must be used if biopsied embryos have to be frozen.

Animals↗
PubMed1994

5-Azacytidine changes gene expression and causes developmental arrest of early chick embryo.

Methylation of DNA appears to be an important maintenance mechanism for inhibiting gene expression during development in vertebrates. 5-azacytidine (5-azaC) is used as an experimental tool for demethylation and it induces differentiation in various systems. In the chick embryo, the first cellular migrations signal the onset of primitive streak and gastrula formation and result in neural induction and morphogenesis of the embryonic axis. In the present work with the early chick embryo, 5-azaC perturbs normal cellular migrations and the embryos produce an atypical short, thickened primitive streak. These embryos have the tendency to form neural tissue but the embryonic axis shows sparse identity of patterning along its length. A small percentage of embryos display formation of double embryonic axes. Blastula embryos show reduced expression of some polypeptides and express characteristic polypeptides which are not present in morula embryos normally. Under the influence of 5-azaC, blastula embryos expressed all the polypeptides which are characteristic of embryos at both the morula and the blastula stages. If 5-azaC perturbs DNA methylation as the chick embryo develops from the histologically simple blastula, then the wave of methylation which has been reported to start at the late blastula and continues during postgastrulation in vertebrate embryos does not seem to be important for the induction of mesodermal and of neural tissues, but is important for the patterning of these tissues.

Animals↗