Proceedings of the XLV Italian Society of Agricultural Genetics - SIGA Annual Congress

Salsomaggiore Terme, Italy - 26/29 September, 2001

ISBN 88-900622-1-5

Oral Communication Abstract

BIOTECHNOLOGIES AND GENETIC MECHANISMS REGULATING THE REPRODUCTION OF DOMESTIC ANIMALS

GANDOLFI F., BREVINI T.A.L., MODINA S.

Istituto di Anatomia degli Animali Domestici, Via Trentacoste 2, 20134 Milano, Italy

fulvio.gandolfi@unimi.it

The application to farm animals of methods for assisted reproduction and genetic modification has been constantly progressing in recent years. This presentation will summarise these methods and their application to various aspects of animal production. Particular emphasis will be given to the most recent and promising developments.

In vitro production of embryos (IVP) is the base for all the other methods aiming at manipulating reproduction or modificating the genome of these species. Oocyte maturation, fertilization and pre-implantation development in vitro, initially limited mainly to ruminants, can now be achieved also in pigs, horses and carnivores, though with different degrees of efficiency. IVP together with cryopreservation and ultrasound-guided follicle aspiration can be applied to breeding schemes for improving selection intensity and efficiency.

The easy access to a large number of zygotes at a reasonable cost, provided by IVP, has been used for inserting transgenes in the genome of ruminants and pigs. However the very low efficicency of all available methods still limits severly any attempt of commercial exploitation. For this reason the develoment of an easy and efficient system for inserting exogenous genes in farm animals is a prime research goal. To this pourpose the use of spermatozoa as transgene vectors has been extensively investigated with alternate results that recently seem to become promising.

Originally developed in farm animals, cloning has rapidly become the most applied method for manipulating the genome of these species. Its efficiency, initially as low as that of trasgenesis, is slowly but steadily increasing, though no real progress have so far been done in our understanding of the mechanisms that regulate nuclear reprogramming. Applications of cloning ranges from the propagation of prized livestock to the production of trangenic animals; from the preservation of endangered species to the generation of stem cells for tissue transplantation and/or gene therapy. Undoubtedly, many more applications and benefits are yet to be imagined.

In order to be able to achieve all the potential benefits that reproductive biotechnologies hold for us it is necessary to understand the mechanisms that regulate early embryonic development.

In our laboratory we have characterized cloned and sequenced the bovine homologue of the gene Oct-4 a transcription factor belonging to the POU family. Its expression, in mouse, is restricted to totipotent and pluripotent cell lineages therefore it was an interesting candidate marker for characterizing putative stem cells and monitoring nuclear reprogramming. Results indicated that bovine Oct-4 has a specific expression profile in bovine and pig pre-implantation embryos that is different from mouse. However Oct-4 expression pattern was identical between in vitro generated embryos and embryos generated by transplantation of a granulosa cell nucleus in an enucleated bovine oocyte.

Beside the analysis of single genes we have also studied the molecular mechanisms of the oocyte competence to be fertilized and sustain embryonic development. Our work indicated that maternal mRNA stored in the ooplasm plays an important role. Molecules of mRNA are synthetised during oogenesis in order to be utilised during the first cleavages before the embryonic genome is activated and endogenous transcription begins. In cattle the maternal to embryonic transition takes place after the third cleavge, and maternal mRNA transcription is regulated by the extension of the poly(A) tail at the 3’ end. We demonstrated that any event with a negative impact on oocyte developmental competence has also the capability to alter the polyadenylation level of some maternal RNA molecules.