resurrection of a bull by cloning from organs frozen without cryoprotectant in a ？80°c freezer for a decade复活一头公牛的克隆器官没有冷冻保护剂在冻结 80°c冷冻了十年.pdfVIP

Resurrection of a Bull by Cloning from Organs Frozen without Cryoprotectant in a 280uC Freezer for a Decade Yoichiro Hoshino1,3, Noboru Hayashi1,2,3, Shunji Taniguchi2,3, Naohiko Kobayashi1,2,3, Kenji Sakai1,2, Tsuyoshi Otani1,2, Akira Iritani2,3, Kazuhiro Saeki2,3* 1 Gifu Prefectural Livestock Research Institute, Takayama, Gifu, Japan, 2 Department of Genetic Engineering, Kinki University, Kinokawa Wakayama, Japan, 3 Wakayama Industry Promotion Foundation, Wakayama, Japan Abstract Frozen animal tissues without cryoprotectant have been thought to be inappropriate for use as a nuclear donor for somatic cell nuclear transfer (SCNT). We report the cloning of a bull using cells retrieved from testicles that had been taken from a dead animal and frozen without cryoprotectant in a 280uC freezer for 10 years. We obtained live cells from defrosted pieces of the spermatic cords of frozen testicles. The cells proliferated actively in culture and were apparently normal. We transferred 16 SCNT embryos from these cells into 16 synchronized recipient animals. We obtained five pregnancies and four cloned calves developed to term. Our results indicate that complete genome sets are maintained in mammalian organs even after long-term frozen-storage without cryoprotectant, and that live clones can be produced from the recovered cells. Citation: Hoshino Y, Hayashi N, Taniguchi S, Kobayashi N, Sakai K, et al. (2009) Resurrection of a Bull by Cloning from Organs Frozen without Cryoprotectant in a 280uC Freezer for a Decade. PLoS ONE 4(1): e4142. doi:10.1371/journal.pone.0004142 Editor: Hiroaki Matsunami, Duke Unviersity, United States of America Received October 27, 2008; Accepted December 1, 2008; Published January 8, 2009 Copyright: 2009 Hoshino et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits