Two clones produced in the study survived to adulthood Japanese scientists have managed to create clones from the bodies of mice which have been frozen for 16 years. Cloning has largely been done using just live donor cells, transferring their DNA to recipient eggs. Using previously frozen cells runs the risk of ice damage to the DNA unless carefully handled. The scientists in Kobe, Japan, said their technique raised the possibility of recreating extinct creatures, such as mammoth, from their frozen remains. Many of the successful clones since Dolly the sheep was born in 1996 have been created by a method where the nucleus of a cell has been removed, placed in an empty egg and kick-started into replicating by chemicals or electricity. It is not the only cloning technique, and Australian researchers reported cloning a pig in 2001 from cells that had been frozen for two years. The Adelaide-based team said its cloning method differed from the Dolly approach in important respects. Frozen bodies The Japanese research was undertaken at Kobe's Centre for Developmental Biology and is reported in the journal Proceedings of the National Academy of Sciences (PNAS). The work extends the time frozen material can be held before it is used to clone an animal. Pigs have been cloned using frozen material before now The scientists said they created their mice from the brain cells of rodents that had been kept in laboratory conditions at -20C. Obviously, these are ideal conditions - far removed from the uncontrolled conditions of the Siberian steppe where mammoth remains dating back 40,000 years have emerged from the permafrost. DNA previously removed from such animals has been found to be highly degraded. Even if good material were available major obstacles would have to be overcome before such a technique could be used on extinct or even endangered animals - such as finding a suitable species to provide recipient eggs and surrogate mothers. The donor genetic material came from animals stored at -20C for 16 years

Professor John Armitage is director of tissue banking at the Bristol Eye Hospital, UK. He commented: "Mitochondrial and some nuclear DNA fragments have previously been isolated from mammoths frozen in permafrost, but the key question is whether sufficiently intact nuclei could be extracted from mammoth cells, which will have been frozen for at least 10,000 years at relatively high sub-zero temperatures. Professor Armitage added: "-20C, the temperature at which the mice used in these experiments were stored, is insufficient to stop physical and chemical reactions of biological significance - even food in a domestic freezer has a limited storage time based on changing texture and taste. "To achieve long-term storage of viable cells, including embryos, requires far lower temperatures of at least -140C in the presence of cryoprotectants." It is conceivable the techniques being developed might have some application in future stem cell therapies in humans where a cloning process was used to generate particular tissues for transplantation. Viable eggs, sperm and embryos are already retrieved from the frozen state for use in in vitro fertilisation (IVF).



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