The Frozen Zoo® of San Diego Zoo Global: Past, Present, and Future

Dr. Oliver Ryder, Barbara Durrant, Marlys L. Houck, Marisa Korody, Cynthia Steiner, Institute for Conservation Research, San Diego Zoo Global

Cryobanking of viable tissue culture cells at the San Diego Zoo, named the “Frozen Zoo® by its founder, Kurt Benirschke, has contributed knowledge advancing conservation science and species recovery from its beginnings in 1975. This diverse collection of cryopreserved diploid fibroblast cell cultures contributes to the legacy his endeavor. Banked gametes and cryopreserved reproductive tissues expanded the possibility of conservation applications. Identification of chromosomal errors affecting fertility of mammalian species as diverse as tigers, dik-diks, and gorillas has been enabled by its collections. Key studies delimiting species boundaries, evolutionary relationships, phylogeny and systematics of mammals and birds have utilized samples from the Frozen Zoo and its extended network of samples. Major contributions to the advancement of comparative vertebrate genomics have been facilitated by the collections of the Frozen Zoo. A crucial contributor to the 200 Mammals project, Genome10K, and the Vertebrate genome project, intact cells and high molecular weight DNA extracts from the Frozen Zoo have facilitated whole genome sequencing efforts and, notably, high quality de novo genome assemblies. Cryobanking of viable early passage diploid fibroblasts has allowed investigation of somatic cell nuclear transfer cloning technology to be pursued in the conservation context. Live offspring of two endangered species of wild cattle, the gaur, and the Javan banteng have been produced with cells banked for decades in the Frozen Zoo. Fibroblast cells from the Frozen Zoo were used for the first reported cellular reprogramming of endangered species to produce induced pluripotent stem cells (iPSC), including an African primate, the drill, and the critically endangered northern white rhinoceros were successfully reprogrammed. These studies have been extended and recently, using nonintegrating methods, iPSC have from two southern white and eight northern white rhinoceroses have been produced and characterized, an effort crucial to preventing its otherwise-certain extinction.