Cloning extinct species #1: A how-to guide

By Hilary Miller 30/04/2010

 Fancy seeing herds of mammoths running across the tundra, moa crashing through the undergrowth, or perhaps a tasmanian tiger lurking in the Aussie bush? Well in the near future these images might not just be the stuff of far-fetched Hollywood movie plots.  Advances in molecular biology and genomics mean that the ability to clone extinct species is getting closer. In theory, at least.  In this post I’m going to look at how one would go about bringing back to life their favourite extinct species, and in a later post I’ll discuss whether we should bother.

The technology required to clone extinct species is largely already here.  In fact, in November 2008 Japanese researchers reported cloning mice that had been frozen 16 years, and last year researchers in Spain reported cloning the bucardo (Pyrenean ibex), a subspecies of the Spanish ibex which went extinct in 2000, from a piece of frozen skin (although the animal died soon after birth).  Whole genome sequencing for extinct species is also becoming a reality – a partial genome sequence for the mammoth was published in Nature in 2008.   

The Woolly Mammoth may have had its genome sequenced, but cloning is still a long way off

While a genome sequence is a good start for resurrecting an extinct species, that alone is not enough.  In order to turn the DNA sequence of the genome into a living organism, the DNA has to be packaged into chromosomes and the information in the DNA expressed in living cells.  Conventional reproductive cloning (like that used to create Dolly the sheep) is performed using Somatic Cell Nuclear Transfer (SCNT), where the genetic material (ie the nucleus of a cell) from one organism is transferred into an egg from which the nucleus has been removed.  Because the nucleus that is transferred comes from a somatic cell, it already has a full diploid genome (ie 2 copies of each chromosome), and with a bit of kick-starting behaves like a freshly fertilized egg, developing into an organism identical to the one from which the nucleus came from.   

So to clone an extinct species using this method, you would need a closely related living species to provide an egg, and an intact nucleus containing a full set of chromosomes in good condition.  This is where things get tricky, as the DNA extracted from remains of extinct organisms is normally extremely degraded.  Even obtaining genome sequences from this material is only possible due to the advent of high throughput “next generation” sequencers, which can sequence billions of short fragments of DNA.  Heavy duty computer power is then required to stitch those sequences back together to decipher the genome sequence – a bit like a giant jigsaw puzzle.  Having a genome sequence for a closely related species for comparison helps enormously with this assembly, as it can be used as a sort of template on which to assemble the genome of the extinct species.   

Once you have the genome there are two ways in which (theoretically) you could rebuild the chromosomes of the extinct organism. Firstly, you could modify the DNA of a closely related species at each base that it differs from the extinct species.  For instance in the case of mammoths you could use African elephant DNA, which differs at about 400,000 sites.  Or alternatively, you could synthesize the whole genome from scratch.   However, so far only small bacterial genomes have been successfully synthesized, and then there is the problem of how to package the synthesized DNA into chromosomes and enclose the whole lot in a nuclear membrane.  These are not a trivial problems, as for many extinct species we don’t even know how many chromosomes there were, let alone which bits of DNA go where*.    

Of course these problems of rebuilding a genome can be circumvented if well-preserved tissue from the organism is available. If this is the case, an intact nucleus could be transferred directly a la Dolly the sheep.  This was the method used to clone the bucardo.  In 1999, researchers in Spain had the foresight to take skin samples from the last living bucardo, and froze them in liquid nitrogen.  The bucardo genetic material was then transferred into the eggs of a domestic goat, and the resulting embryo implanted into other species of spanish ibex or goat-ibex hybrids.  Researchers at the Audubon Centre for Research of Endangered Species (part of Audubon Nature Institute) are also using this method to clone critically endangered species such as the african wildcat.    

Using an intact nucleus from frozen tissue is obviously a much easier approach than rebuilding a whole genome from scratch, but for the majority of extinct species we don’t have a good enough tissue source.  Even exceptionally well preserved remains, such as those frozen in permafrost, show significant amounts of degradation, and the chances of finding an intact nucleus with undegraded DNA in a specimen that is several thousand years old are virtually nil.  Organisations around the world are now beginning to realise the importance of frozen tissue resources, and are creating frozen zoos, where archives of tissue from endangered species are cryopreserved for future use.  These resources won’t help resurrect the mammoth, but might facilitate future cloning endeavours on critically endangered species.  

Its also unlikely we’ll be seeing cloned moa, huia or giant eagles any time soon, as somatic cell nuclear transfer currently only works in mammals.  No one has yet successfully cloned a bird, reptile, amphibian or fish using these methods, because differences in the structure of the eggs in these species provide added complications.  Oh and you can forget about cloning dinosaurs too.  Although researchers at North Carolina State University have claimed to have extracted protein from dinosaur bones, no DNA has been found and it seems extremely unlikely that DNA could survive intact for more than a few hundred thousand years.  Virtually all reports of DNA extracted from samples that are many millions of years old have failed closer inspection, so it seems that the idea of extracting DNA from dinosaur blood found in mosquitos fossilized in amber belongs firmly in the realm of fantasy.  

* This news feature in Nature provides more detail and a really nice explanation of the methodological difficulties that have yet to be overcome with this type of cloning