Full-term development has now been achieved in several mammalian species by transfer of somatic nuclei into enucleated oocytes [1, 2]. Although a high proportion of such reconstructed embryos can evolve until the blastocyst stage, only a few percent develop into live offspring, which often exhibit developmental abnormalities [3, 4]. Regulatory epigenetic markers such as DNA methylation are imposed on embryonic cells as normal development proceeds, creating differentiated cell states. Cloned embryos require the erasure of their somatic epigenetic markers so as to regain a totipotent state [5]. Here we report on differences in the dynamics of chromosome methylation between cloned and normal bovine embryos before implantation. We show that cloned embryos fail to reproduce distinguishable parental-chromosome methylation patterns after fusion and maintain their somatic pattern during subsequent stages, mainly by a highly reduced efficiency of the passive demethylation process. Surprisingly, chromosomes appear constantly undermethylated on euchromatin in morulae and blastocysts, while centromeric heterochromatin remains more methylated than that of normal embryos. We propose that the abnormal time-dependent methylation events spanning the preimplantation development of clones may significantly interfere with the epigenetic reprogramming, contributing to the high incidence of physiological anomalies occurring later during pregnancy or after clone birth.

-Assisted reproductive techniques (gamete cryopreservation, artificial insemination, embryo transfer, and in vitro fertilization) allow to propagate small fragmented populations of wild endangered species or domestic breeds. There are the best way for producing several offspring from selected genitors in order to avoid inbreeding depression. However, few mammalian species have been well studied for their reproductive biology whereas huge differences have been observed between these species. Furthermore, materials, methods and experimental designs have to be adapted for each case and each limiting factor (wildness, poor quantity of biological material, disparate locations). Genome resource banking is currently arising and the most applied reproductive biotechnology remains artificial insemination. Assisted reproductive techniques currently developed in domestic species (intracytoplasmic sperm injection, nuclear transfer) may offer new opportunities for the propagation of endangered species. endangered species / artificial insemination / cryopreservation / in vitro fertilization / embryo Résumé -Biotechnologies de la reproduction appliquées aux mammifères en voie de disparition. Les techniques de reproduction assistée (cryoconservation des gamètes, insémination artificielle, transfert embryonnaire, fécondation in vitro) permettent d'accroître des populations, parfois dispersées géographiquement, d'espèces sauvages ou domestiques en voie de disparition. Ces méthodes sont le meilleur moyen pour produire plusieurs descendants à partir de géniteurs sélectionnés de façon à éviter la consanguinité. La biologie de la reproduction est connue pour peu d'espèces de mammifères alors que de grandes différences ont pourtant été mises en évidence entre ces espèces. En outre, les matériels, les méthodes, ainsi que les schémas expérimentaux doivent être adaptés à chaque cas et pour chaque facteur limitant (animaux sauvages, peu de matériel biologique, populations dispersées). Des banques génétiques sont actuellement en voie de constitution mais la technique la plus utilisée reste l'insémination artificielle. Cependant, des biotechnologies de la reproduction en cours de mise au point chez les animaux domestiques (injection intra-cytoplasmique de spermatozoïde, transfert nucléaire) pourraient offrir de nouvelles possibilités pour l'accroissement de populations menacées. espèces menacées / insémination artificielle / congélation / fécondation in vitro / embryon Reprod. Nutr. Dev. 40 (2000)

Abstract. Sustaining viable populations of all wildlife species requires the maintenance of habitat, as well as an understanding of the behaviour and physiology of individual species. Despite substantial efforts, there are thousands of species threatened by extinction, often because of complex factors related to politics, social and environmental conditions and economic needs. When species become critically endangered, ex situ recovery programmes that include reproductive scientists are the usual first line of defence. Despite the potential of reproductive technologies for rapidly increasing numbers in such small populations, there are few examples of success. This is not the result of a failure on the part of the technologies per se, but rather is due to a lack of knowledge about the fundamental biology of the species in question, information essential for allowing reproductive technologies to be effective in the production of offspring. In addition, modern conservation concepts correctly emphasise the importance of maintaining heterozygosity to sustain genetic vigour, thereby limiting the practical usefulness of some procedures (such as nuclear transfer). However, because of the goal of maintaining all extant gene diversity and because, inevitably, many species are (or will become) 'critically endangered', it is necessary to explore every avenue for a potential contributory role. There are many 'emerging technologies' emanating from the study of livestock and laboratory animals. We predict that a subset of these may have application to the rescue of valuable genes from individual endangered species and eventually to the genetic management of entire populations or species. The present paper reviews the potential candidate techniques and their potential value (and limitations) to the study and conservation of rare wildlife species.