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Reproduction, Fertility and Development Reproduction, Fertility and Development Society
Vertebrate reproductive science and technology
RESEARCH ARTICLE

State-of-the-art production, conservation and transfer of in-vitro-produced embryos in small ruminants

Yves Cognié A , Nati Poulin A , Yann Locatelli A and Pascal Mermillod A B
+ Author Affiliations
- Author Affiliations

A INRA, Physiologie de la Reproduction et des Comportements, 37380 Nouzilly, France.

B To whom correspondence should be addressed. email: mermillo@tours.inra.fr

Reproduction, Fertility and Development 16(4) 437-445 https://doi.org/10.1071/RD04029
Accepted: 23 March 2004   Published: 6 July 2004

Abstract

Today, although not efficient enough to replace multiple ovulation and embryo transfer, in vitro embryo production for small ruminants is a platform for new reproductive technologies, such as embryo sexing, transgenesis and cloning. The in vitro embryo-production system developed for sheep and goats is more efficient now than 15 years ago, but could still be improved. Laparoscopic collection of oocytes in live animals treated with gonadotrophin indicates a promising future for the application of this technology to genetic improvement programmes. Oocyte maturation in defined medium with epidermal growth factor and cysteamine appears as efficient as oocyte maturation in follicular fluid-supplemented medium and allows future study of the effect of other factors involved in the cytoplasmic maturation of oocytes from these species. Further efforts have to be made to standardise the semen-capacitating process and to improve the quality and freezability of in-vitro-produced (IVP) embryos. The optimisation of IVP procedures for deer species has required the study of the seasonal variation of oocyte competence and the development of a specific methodology to allow the culture of embryos up to the blastocyst stage.

Extra keywords: blastocyst, caprine, deer, development, fertilisation, freezing, maturation, oocyte, ovine.


References

Abeydeera, L. R. , Wang, W. H. , Cantley, T. C. , Prather, R. S. , and Day, B. N. (1998). Presence of β-mercapto-ethanol can increase the glutathione content of pig oocytes matured in vitro and the rate of blastocyst development after in vitro fertilization. Theriogenology 50, 747–756.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Accardo, C. , Dattena, M. , Pilichi, S. , Mara, L. , Chessa, B. , and Cappai, P. (2004). Effect of recombinant human FSH and LH on in vitro maturation of sheep oocytes: embryo development and viability. Anim. Reprod. Sci. 81, 77–86.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Aguilar, B., Roche, A., Olivera, J., Folch, J., and  Alabart, J. L. (2002). Oocyte retrieval after repeated ovum pick-up in unstimulated sheep and goat. In ‘Proceedings of the 18th Meeting Association Européenne de Transfert Embryonnaire (AETE)’.  pp. 30. (INRA: Nouzilly, France.) [Abstract]

Alberio, R. , Echegoyen, E. , Olivera, J. , Roche, A. , Alabart, J. , Fernandez-Arias, A. , and Folch, J. (2001). Performance in an OPU program of FSH stimulated or non stimulated Romanov and Rasa Aragonesa ewe breeds. Theriogenology 55, 509.[Abstract]


Baldassarre, H. , Furnus, C. C. , de Matos, D. G. , and Pessi, H. (1996). In vitro production of sheep embryos using laparoscopic folliculocentesis: alternative gonadotrophin treatments for stimulation of oocyte donors. Theriogenology 45, 707–717.
Crossref | GoogleScholarGoogle Scholar |

Baldassarre, H. , Wang, B. , Kafidi, N. , Keefer, C. , Lazaris, A. , and Karatzas, C. N. (2002). Advances in the production and propagation of transgenic goats using laparoscopic ovum pick-up and in vitro embryo production technologies. Theriogenology 57, 275–284.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Baldassarre, H. , Wang, B. , Kafidi, N. , Gauthier, M. , and Neveu, N. (2003). Production of transgenic goats by pronuclear microinjection of in vitro produced zygotes derived from oocytes recovered by laparoscopy. Theriogenology 59, 831–839.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Berg, D. K. , and Asher, G. W. (2003). New developments reproductive technologies in deer. Theriogenology 59, 189–205.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Berg, D. K. , Thompson, J. G. , and Asher, G. W. (2002). Development of in vitro embryo production systems for red deer (Cervus elaphus). 3. In vitro fertilization using sheep serum as a capacitating agent and the subsequent birth of calves. Anim. Reprod. Sci. 70, 85–98.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Blash, S. , Melican, D. , and Gavin, W. (2000). Cryopreservation of epididymal sperm obtained at necropsy from goats. Theriogenology 54, 899–905.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Bols, P. E. J. , Van Soom, A. , Ysebaert, M. T. , Vandenheede, J. M. M. , and de Kruif, A. (1996). Effects of aspiration vacuum and needle diameter on cumulus oocyte complex morphology and developmental capacity of bovine oocytes. Theriogenology 45, 1001–1014.
Crossref | GoogleScholarGoogle Scholar |

Byrne, G. P. , Lonergan, P. , Wade, M. , Duffy, P. , Donovan, A. , Hanrahan, J. P. , and Boland, M. P. (2000). Effect of freezing rate of ram spermatozoa on subsequent fertility in vivo and in vitro.  Anim. Reprod. Sci. 62, 265–275.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Charpigny, G., Marquant-Leguienne, B., Abd el Razek, I., Humblot, P., and  Mermillod, P. (2000). Differences in fatty acids incorporation into lipids between in vivo and in vitro produced embryos from ruminant species. In ‘Proceedings of the 7th International Conference on Goats, Tours, France’. (Eds. L Gruner and Y Chebert) (Institut de l'Élevage and INRA: Paris, France.) Vol. 2. [Abstract]

Cognié, Y. (1999). State of the art in sheep–goat embryo transfer. Theriogenology 51, 105–116.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Cognié, Y., and  Poulin, N. (2000). Developmental competence of goat oocytes is increased after in vitro maturation with follicular fluid from goats stimulated by gonadotropins. In ‘Proceedings of the 14th International Congress on Animal Reproduction’.  pp. 39. (ICAR: Stockholm, Sweden.) Vol. 18. [Abstract]

Cognié, Y. , Poulin, N. , Pisselet, C. , and Monniaux, D. (1995). Effect of atresia on the ability of follicular fluid to support cytoplasmic maturation of sheep oocytes in vitro.  Theriogenology 43, 188.
Crossref | GoogleScholarGoogle Scholar |

Cognié, Y., Poulin, N., Baril, G., Guignot, F., Beckers, J. F., and  Mermillod, P. (2001). Embryo survival after transfer of in vitro and in vivo produced goat embryos. In ‘Proceedings of the 17th Meeting Association Européenne de Transfert Embryonnaire (AETE)’.  pp. 110. (INRA: Nouzilly, France.) [Abstract]

Cognié, Y., Poulin, N., and  Mermillod, P. (2002). Cysteamine improves in vitro goat oocyte maturation in defined medium. In ‘Proceedings of the 18th Meeting Association Européenne de Transfert Embryonnaire (AETE)’.  pp. 154. (INRA: Nouzilly, France.) [Abstract]

Cognié, Y. , Baril, G. , Poulin, N. , and Mermillod, P. (2003). Current status of embryo technologies in sheep and goat. Theriogenology 59, 171–188.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Comizzoli, P. , Mermillod, P. , Cognié, Y. , Chai, N. , Legendre, X. , and Mauget, R. (2001). Successful in vitro production of embryos in the red deer (Cervus elaphus) and the sika deer (Cervus nippon). Theriogenology 55, 649–659.
Crossref | GoogleScholarGoogle Scholar | PubMed |

De Matos, D. G. , Furnus, C. C. , Moses, D. F. , and Baldassarre, H. (1995). Effect of cystamine on glutathione level and development capacity of bovine oocytes matured in vitro.  Mol. Reprod. Dev. 42, 432–436.
PubMed |

De Matos, D. G. , Gasparini, B. , Pascalini, S. R. , and Thompson, J. G. (2002). Effect of glutathione synthesis during in vitro maturation of ovine oocytes on embryo development and intracellular peroxide content. Theriogenology 57, 1443–1451.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Dieleman, S. J. , Hendriksen, P. J. M. , Viuff, D. , Thomsen, P. D. , and Hyttel, P. , et al. (2002). Effects of in vivo prematuration and in vivo final maturation on developmental capacity and quality of pre-implantation embryos. Theriogenology 57, 5–20.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Duszewska, A. M. , Reklewski, Z. , Pienkowski, M. , Karasiewicz, J. , and Modlinski, J. A. (2000). Development of bovine embryos on Vero/BRL cell monolayers (mixed co-culture). Theriogenology 54, 1239–1247.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Gandolfi, F. (1994). Autocrine, paracrine, and environmental factors influencing embryonic development from zygote to blastocyst. Theriogenology 41, 95–100.


Gandolfi, F. , and Moor, R. M. (1987). Stimulation of early embryonic development in the sheep by co-culture with oviduct epithelial cells. J. Reprod. Fertil. 81, 23–28.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Gardner, D. K. , and Lane, M. (1998). Culture of viable human blastocysts in defined sequential serum-free media. Hum. Reprod. 13((Suppl. 3)), 148–159.
Crossref | GoogleScholarGoogle Scholar | PubMed | PubMed |

Gardner, D. K. , Lane, M. , Spitzer, A. , and Batt, A. (1994). Enhanced rates of cleavage and development for sheep zygotes cultured to the blastocyst stage in vitro in the absence of serum and somatic cells: amino acids, vitamins, and culturing embryos in groups stimulate development. Biol. Reprod. 50, 390–400.
PubMed |

Gianaroli, M. C. , Magli, M. C. , Ferraretti, A. P. , Fiorentino, A. , Tosti, E. , and Panzella, S. (1996). Reducing the time of sperm–oocyte interaction in human in vitro fertilization improves the implantation rate. Hum. Reprod. 11, 166–171.
PubMed |

Graff, K. J. , Meintjes, M. , Dyer, V. W. , Paul, J. B. , Denniston, R. S. , Ziomeck, C. , and Godke, R. A. (1999). Transvaginal ultrasound-guided oocyte retrieval following FSH stimulation of domestic goats. Theriogenology 51, 1099–1119.
PubMed |

Guérin, Y., Cognié, Y., and  Poulin, N. (1992). Fertilizability of freshly ejaculated or frozen ram semen in vitro. In ‘Proceedings of the 12th International Congress on Animal Reproduction’.  pp. 1418–1420. (ICAR: La Haye, The Netherlands.) Vol. 3.

Guler, A. , Poulin, N. , Mermillod, P. , Terqui, M. , and Cognié, Y. (2000). Effect of growth factors, EGF and IGF-I, and estradiol on in vitro maturation of sheep oocytes. Theriogenology 54, 209–218.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Heyman, Y. , Lavergne, Y. , Charpigny, G. , Renard, J. P. , and Ménézo, Y. (2002). Development of bovine embryos cultured in the serum free sequential media ISM1/ISM2. Theriogenology 57, 671. [.


Holm, P. , Walker, S. K. , Petersen, B. A. , Ashman, R. J. , and Seamark, R. F. (1994). In vitro vs in vivo culture of ovine IVM/IVF ova: effect on lambing. Theriogenology 41, 217. [.


Holm, P. , Walker, W. H. , and Seamark, R. F. (1996). Embryo viability, duration of gestation and birth weight in sheep after transfer of in vitro matured and in vitro fertilized zygotes cultured in vitro or in vivo.  J. Reprod. Fertil. 107, 175–181.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Huneau, D. , Crozet, N. , and Ahmed-Ali, M. (1994). Estrous sheep serum as a potent agent for ovine IVF: effect on cholesterol efflux from spermatozoa and the acrosome reaction. Theriogenology 42, 1017–1028.
Crossref | GoogleScholarGoogle Scholar |

Izadyar, F. , Van Tol, H. T. A. , Colenbrander, B. , and Bevers, M. M. (1997). Stimulatory effect of growth hormone on in vitro maturation of bovine oocytes is exerted through cumulus cells and not mediated by IGF-I. Mol. Reprod. Dev. 47, 175–180.
PubMed |

Izquierdo, D. , Villamediana, P. , Lopez-Bejar, M. , and Paramio, M. T. (2002). Effect of in vitro and in vivo culture on embryo development from prepubertal goat IVM/IVP oocytes. Theriogenology 57, 1431–1441.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Kelly, J. M. , Kleeman, D. O. , Kuwayama, M. , and Walker, S. K. (2004). Vitrification of in vitro-produced bovine and ovine embryos using the minimum volume cooling cryotop method.  Reprod. Fertil. Dev. 16, 172.
Crossref | GoogleScholarGoogle Scholar |

Kochhar, H. P. S. , Buckrell, B. C. , Pollard, J. W. , and King, W. A. (1999). Cleavage, sex and cell number of ovine embryos developed after different sperm-oocyte co-incubation intervals. Theriogenology 51, 322. [.


Kuhholzer, B. , Muller, S. , Treuer, A. , Seregi, J. , Besenfelder, U. , and Brem, G. (1997). Repeated endoscopic ovum pick-up in hormonally untreated ewes: a new technique. Theriogenology 48, 545–550.
Crossref | GoogleScholarGoogle Scholar |

Langlais, J. , Kan, F. W. K. , Granger, L. , Raymond, L. , Bleau, G. , and Roberts, K. D. (1988). Identification of sterol acceptors that stimulate cholesterol efflux from human spermatozoa during in vitro capacitation. Gamete Res. 20, 185–201.
PubMed |

Locatelli, Y. , Poulin, N. , Baril, G. , Touzé, J.-L. , Bouttier, A. , Cognié, Y. , and Mermillod, P. (2004a). FSH treatment before laparoscopic ovum pick-up in goat influences quantity and quality of recovered oocytes. Reprod. Fertil. Dev. 16,


Locatelli, Y., Vallet, J. C., Huyghe, F., Legendre, X., Cognié, Y., and  Mermillod, P. (2004b). Repeated stimulations of follicular growth, ovum pick-up and in vitro production of embryos during seasonal anoestrus and oestrus in sika deer (Cervus nippon nippon). In ‘Proceedings of the 15th International Congress of Animal Reproduction, Porto Seguro, Brazil, August 2004’. (Brazilian College of Animal Reproduction: Belo-Horizonte, Brazil.) [Abstract]

Lonergan, P. , Khatir, H. , Piumi, F. , Rieger, D. , Humblot, P. , and Boland, M. P. (1999). Effect of time interval from insemination to first cleavage on the developmental characteristics, sex and pregnancy rates following transfer of bovine preimplantation embryos. J. Reprod. Fertil. 117, 159–167.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Menck, M. C. , Guyader-Joly, C. , Peynot, N. , Le Bourhis, D. , Lobo, R. B. , Renard, J. P. , and Heyman, Y. (1997). Beneficial effects of Vero cells for developing IVF bovine eggs in two different coculture systems. Reprod. Nutr. Dev. 37, 141–150.
PubMed |

Mermillod, P. , Oussaid, B. , and Cognié, Y. (1999). Aspects of follicular and oocyte maturation that affect the developmental potential of embryos. J. Reprod. Fertil. Suppl. 54, 449–460.
PubMed |

Merton, J. S. , de Roos, A. P. , Mollaart, E. , de Ruigh, L. , Kaal, L. , Vos, P. L. , and Dieleman, S. J. (2003). Factors affecting oocyte quality and quantity in commercial application of embryo technologies in the cattle breeding industry. Theriogenology 59, 651–674.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Moor, R. M. (1978). Role of steroids in the maturation of ovine oocytes. Ann. Biol. Anim. Biochim. Biophys. 18, 477–482.


Moor, R. M. , and Trounson, A. O. (1977). Hormonal and follicular factors affecting maturation of sheep oocytes in vitro and their subsequent development capacity. J. Reprod. Fertil. 49, 101–109.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Murdoch, W. J. , and Van Kirk, E. A. (2001). Estrogenic upregulation of DNA polymerase β in oocytes of preovulatory follicles. Mol. Reprod. Dev. 58, 417–423.
Crossref | GoogleScholarGoogle Scholar | PubMed |

O’Brien, J. K. , Hollinshead, F. K. , and Maxwell, W. M. C. (2004). In vivo development capacity of in vitro-produced embryos derived from sex-sorted and re-cryopreserved frozen–thawed ram sperm. Reprod. Fertil. Dev. 16, 286.
Crossref | GoogleScholarGoogle Scholar |

Oussaid, B. , Mariana, J.-C. , Poulin, N. , Fontaine, J. , Lonergan, P. , Beckers, J.-F. , and Cognié, Y. (1999). Reduction of the developmental competence of sheep oocytes by inhibition of LH pulses during the follicular phase with a GnRH antagonist. J. Reprod. Fertil. 117, 71–77.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Ptak, G. , Dattena, M. , Loi, P. , Tischner, M. , and Cappai, P. (1999). Ovum pick-up in sheep: efficiency of in vitro embryo production, vitrification and birth of offspring. Theriogenology 52, 1105–1114.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Ranilla, M. J. , Gebbie, F. E. , King, M. E. , Carolan, C. , Sinclair, K. D. , Watt, R. G. , and Dolman, D. F. (1998). The incidence of embryo and fetal loss following the transfer of in vitro-cultured sheep embryos. Theriogenology 49, 248.
Crossref | GoogleScholarGoogle Scholar |

Rho, G. J. , Hahnel, A. C. , and Betteridge, K. J. (2001). Comparisons of oocyte maturation times and of three methods of sperm preparation for their effects on the production of goat embryos in vitro.  Theriogenology 56, 503–516.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Rizos, D. , Ward, F. , Duffy, P. , Boland, P. M. , and Lonergan, P. (2002a). Consequences of bovine oocyte maturation, fertilization or early embryo development in vitro versus in vivo: implications for blastocyst yield and blastocyst quality. Mol. Reprod. Dev. 61, 234–248.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Rizos, D. , Fair, T. , Papadopoulos, S. , Boland, M. P. , and Lonergan, P. (2002b). Developmental, qualitative, and ultrastructural differences between ovine and bovine embryos produced in vivo or in vitro.  Mol. Reprod. Dev. 62, 320–327.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Rodriguez-Gonzalez, E. , Lopez-Bejar, M. , Mertens, M. J. , and Paramio, M. T. (2003). Effects on in vitro embryo development and intracellular glutathione content of the presence of thiol compounds during maturation of prepubertal goat oocytes. Mol. Reprod. Dev. 65, 446–453.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Silva, C. C. , Groome, N. P. , and Knight, P. G. (1999). Demonstration of a suppressive effect of inhibin α subunit on the developmental competence of in vitro matured bovine oocytes. J. Reprod. Fertil. 115, 381–388.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Sinclair, K. D. , McEvoy, T. G. , Maxfield, E. K. , Maltin, C. A. , Young, L. E. , Wilmut, I. , Broadbent, P. J. , and Robinson, J. J. (1999). Aberrant fetal growth and development after in vitro culture of sheep zygotes. J. Reprod. Fertil. 116, 177–186.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Soler, A. J. , Garcia, A. J. , Fernandez-Santos, M. J. , Esteso, M. C. , and Garde, J. J. (2003). Effects of thawing procedure on postthawed in vitro viability and in vivo fertility of red deer epididymal spermatozoa cryopreserved at –196°C. J. Androl. 24, 746–756.
PubMed |

Stock, A. E. , Woodruff, T. K. , and Smith, L. C. (1997). Effects of inhibin A and activin A during in vitro maturation of bovine oocytes in hormone- and serum free medium. Biol. Reprod. 56, 1559–1564.
PubMed |

Sumantri, C. , Boediono, A. , Ooe, M. , Murakami, M. , Saha, S. , and Susuki, T. (1997). The effect of sperm exposure time on in vitro development using sperm from a tetraparental chimeric bull. Anim. Reprod. Sci. 48, 187–195.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Tervit, H. R. (1996). Laparoscopy/laparotomy oocyte recovery and juvenile breeding. Anim. Reprod. Sci. 42, 227–238.
Crossref | GoogleScholarGoogle Scholar |

Tervit, H. R., Smith, J. F., McGowan, L. T., Wells, R. W., and  Pugh, P. A. (1992). Laparoscopic recovery of ovarian oocytes from slaughtered or living sheep. In ‘Proceedings of the Australian Society for Reproductive Biology’.  pp. 2. (Australian Society for Reproductive Biology: Canberra, Australia.)

Tervit, H. R. , Pugh, P. A. , McGowan, L. T. , Bell, A. C. S. , and Wells, R. W. (1994). The freezability of sheep embryos is affected by culture system and source (in vivo or in vitro derived). Theriogenology 41, 315.


Thompson, J. G. (2000). In vitro culture and embryo metabolism of cattle and sheep embryos: a decade of achievement. Anim. Reprod. Sci. 60–61, 263–275.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Traldi, A. S. , Leboeuf, B. , Cognié, Y. , Poulin, N. , and Mermillod, P. (1999). Comparative results of in vitro and in vivo survival of vitrified in vitro produced goat and sheep embryos. Theriogenology 51, 175.


Vanroose, G. , Van Soom, A. , and de Kruif, A. (2001). From co-culture to defined medium: state of the art and practical considerations. Reprod. Domest. Anim. 36, 25–28.
Crossref | GoogleScholarGoogle Scholar | PubMed | PubMed |

Walker, S. K. , Hartwitch, K. M. , and Seamark, R. F. (1996). The production of unusually large offspring following embryo manipulation: concepts and challenges. Theriogenology 45, 111–120.
Crossref | GoogleScholarGoogle Scholar |

Wang, B. , Baldassarre, H. , Tao, T. , Gauthier, M. , and Neveu, N. , et al. (2002). Transgenic goats produced by DNA pronuclear microinjection of in vitro derived zygotes. Mol. Reprod. Dev. 63, 437–443.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Wang, B. , Baldassarre, H. , Pierson, J. , Cote, F. , Rao, K. M. , and Karatzas, C. N. (2003). The in vitro and in vivo development of goat embryos produced by intracytoplasmic sperm injection using tail-cut spermatozoa. Zygote 11, 219–227.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Ward, F. , Enright, B. , Rizos, D. , Boland, M. , and Lonergan, P. (2002). Optimization of in vitro bovine embryo production: effect of duration of maturation, length of gamete co-incubation, sperm concentration and sire. Theriogenology 57, 2105–2117.
Crossref | GoogleScholarGoogle Scholar | PubMed |

Watson, A. J. , Watson, P. H. , Warnes, D. , Walker, S. K. , Armstrong, D. T. , and Seamark, R. F. (1994). Preimplantation development of in vitro-fertilized ovine zygote: comparison between coculture on oviduct epithelial cell monolayers and culture under low oxygen atmosphere. Biol. Reprod. 50, 715–724.
PubMed |

Zuelke, K. A. , Jeffay, S. C. , Zucker, R. M. , and Perreault, S. D. (2003). Glutathione (GSH) concentrations vary with the cell cycle in maturing hamster oocytes, zygotes, and pre-implantation stage embryos. Mol. Reprod. Dev. 64, 106–112.
Crossref | GoogleScholarGoogle Scholar | PubMed |