for National Geographic News
Using cells from dead mice frozen for 16 years, a team of Japanese geneticists has successfully created healthy clones of the dead animals.
The breakthrough could pave the way for resurrecting extinct animals, such as the woolly mammoth, from frozen remains, experts say.
"We have demonstrated that even frozen animal tissue can be used to produce clones," said Teruhiko Wakayama, a geneticist at the Riken Center for Developmental Biology in Kobe, Japan.
For their cloning process, Wakayama and his colleagues drew dead brain and blood cells from the frozen mice. The researchers injected the nuclei from the dead cells directly into unfertilized mouse eggs, creating embryos.
It's not known, however, whether nuclei from cells frozen for extended periods of time can be reprogrammed to develop into cloned animals.
So instead of transferring each embryo into a mouse's oviduct (the tube by which eggs leave an ovary), the researchers extracted the inner cell mass from each embryo and generated lines of embryonic stem cells. The researchers created 46 such lines, from which they were able to produce 13 mouse pups.
Embryonic stem cells are pluripotent—capable of becoming many other types of cells.
"These cells are the same as fertilized embryonic stem cells," Wakayama explained.
The scientists then transferred the nuclei from these cells into mouse eggs to produce healthy mouse pups. His findings appear today in the journal Proceedings of the National Academy of Science.
Since this cloning method does not require intact cells from the animal being cloned—cells of frozen animals usually deteriorate—the researchers believe that their technique could now allow them to work on the frozen remains of extinct mammals.
Most other methods rely on a fusion between a cell from the donor (in this case brain and blood cells from the frozen mice) and the egg cell into which the genetic material is inserted, Wakayama explained.
"But in dead cells the cell membrane is broken and the fusion method cannot be used for cloning. In our method, it does not matter whether the donor cells are alive or dead," he added.
This summer, geneticist Jinsong Li of the Shanghai Institutes for Biological Sciences used a two-step process to successfully clone mice from mouse cells that had been frozen for nearly 350 days.
Li said Wakayama and his colleagues have made important progress toward the potential application of nuclear transfer in the cloning of animals frozen under natural conditions.
While the resurrection of extinct species may still be a long way off, Li said that, for the moment, Wakayama's discovery indicates that valuable laboratory and farm animals can be thawed and cloned.
(Related: "Cat Cloning Offered to Pet Owners" [March 25, 2004].)
John Critser is a veterinary pathobiologist at the University of Missouri, Columbia, who has previously transplanted elephant ovarian tissue into mice to produce elephant eggs.
He found the study technically elegant, and added that "it is possible one might find some tissue from some endangered species where the genetic material is preserved well enough to perform [the cloning technique] nuclear transfer, and try to resurrect it."
But he cautions that the frozen remains of extinct species could expose infectious diseases not seen since the animal became extinct. "That adds another level of concern," he added.
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