Protocol

Culturing Tammar Wallaby (Macropus eugenii) Peri-gastrulation Stage Embryos

Department of Zoology, The University of Melbourne, Victoria 3010, Australia

¹Corresponding author (m.renfree{at}unimelb.edu.au).

INTRODUCTION

Tammar wallaby (Macropus eugenii) peri-gastrulation stage embryos can be cultured in complete medium in 95% oxygen for up to 3 d. There is little apparent difference in terms of development between tammar peri-gastrulation stage embryos cultured in DMEM versus Medium 199 with Earle’s salts, or between the supplementation of complete medium with either 10% fetal calf serum (FCS) or 10% wallaby serum (WS). The peri-gastrulation marsupial conceptus is hollow, which means that some techniques, such as grafting, cannot be performed on embryos to be cultured whole because, once punctured, the embryo collapses and does not reinflate. This protocol describes both whole embryo culture and an alternative method of culture. In the latter method, the embryonic disc and some adjoining vascular yolk sac are isolated, laid flat on a membrane, and cultured as a flat mount. Embryos cultured in this way develop to a similar stage as those cultured whole.