A Reliable Procedure for the in Vitro Production of Porcine Embryos
Marlon B. Ocampo1*, Lerma C. Ocampo1, and Rebecca L. Krisher2
1Reproductive Biotechnology Unit, Philippine Carabao Center,
Science City of Munoz, 3120 Nueva Ecija, Philippines
2Department of Animal Science, College of Agriculture,
Consumer and Environmental Sciences,
University of Illinois at Urbana-Champaign, USA
corresponding author: This email address is being protected from spambots. You need JavaScript enabled to view it.
ABSTRACT
Details of the procedures described in this study have been used in our laboratory and are consistent and reproducible in terms of achieving porcine oocyte maturation, its fertilization and production of blastocyst stage embryos. In Experiment 1, both defined and undefined maturation conditions supported nuclear maturation of immature oocytes (more than 80.0%), cleavage rate of around 60.0% and blastocyst formation rate of 40.3% and 23.1%, respectively. In Experiment 2, the resulting cleavage and blastocyst formation rate decreases as the sperm concentration during fertilization increases (from 48.7% to 38.6%). In Experiment 3, both Percoll-gradient and Swim-up method of sperm preparation could be used satisfactorily for fertilization and embryo production in vitro (71.6% vs 72.9% cleavage and 51.2% vs 45.1% blastocyst formation rate, respectively). Also, the use of NCSU-23 medium for embryo culture was effective in supporting the acquisition of early stage porcine embryos the competence to overcome the “cell-block” stage and continue support the development up to the blastocyst stage. The ease and reproducibility of the procedures presented makes it a useful tool for gamete biology research, including the aspects of oocyte maturation, sperm capacitation and acrosome reaction, sperm:egg interaction and associated events, block to polyspermy, understanding various contributing factors to fertilization anomalies and embryo development. Also, the advantages offered by these techniques makes it a suitable alternative for other investigators to do research and apply it in other mammalian species.
INTRODUCTION
In the Philippines, only few researchers have used the in vitro maturation, fertilization and culture (IVM/IVF/IVC) procedures to study the oocyte maturation and its subsequent acquisition of developmental competence, sperm capacitation and acrosome reaction, sperm/egg attachment and penetration, and the eventual development of resulting zygotes to transferable stage in some mammals (Ocampo et al. 1998, 2001a, 2001b; Duran 2004; Atabay et al. 2008, 2010) . . . . . .
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