Genetic scientists have sequenced and analyzed short pieces of DNA preserved in bones from an early Middle Pleistocene horse that had been kept frozen for the last 700,000 years in permafrost at Thistle Creek, Yukon, Canada.

Unlike the small Ice Age horse fossils that are common across the unglaciated areas of the Yukon, Alaska and Siberia that date to the last 100,000 years, the Thistle Creek horse fossil found by Dr Duane Froese from the University of Alberta was at least the size of a modern horse.

The scientists had dated the permafrost at the site from volcanic ashes in the deposits and knew that it was about 700,000 years old – representing some of the oldest known ice in the northern hemisphere. They also knew the fossil was similarly old.

They them extracted collagen from the fossil and found it had preserved blood proteins and that short fragments of ancient DNA were present within the bone.

The DNA showed that the horse fell outside the diversity of all modern and ancient horse DNA ever sequenced consistent with its geologic age. After several years of work, a draft genome of the horse was assembled and is providing new insight into the evolution of horses.

The study, reported in the journal Nature, showed that the horse fell within a line that includes all modern horses and the last remaining truly wild horses, the Przewalski’s Horse from the Mongolian steppes.

The 700,000-year-old horse genome – along with the genome of a 43,000-year-old horse Equus lambei, six present-day horses and a donkey – has allowed the team to estimate how fast mutations accumulate through time. In addition, the new genomes revealed episodes of severe demographic fluctuations in horse populations in phase with major climatic changes.

“Sequencing the first genome from the Middle Pleistocene was by no means straightforward,” said study lead author Dr Ludovic Orlando from the University of Copenhagen.

“This was methodologically challenging but clearly some parameters worked better than others,” said co-author Prof Eske Willerslev, also from the University of Copenhagen.

“Because 700,000 years of evolution and damage, it is not something that does come without any modification to the DNA sequence itself. We had to improve our ability to identify modified and divergent ancient horse sequences by aligning them to the genome of present day horses.”

The scientists also solved a long-standing mystery about the evolutionary origin of Przewalski’s Horse. They revealed that the Przewalski’s Horse population was isolated from the lineage leading to present-day domesticated horses about 50,000 years ago. They also showed that pure Przewalski’s Horse lineages are still living today. This suggests that the Przewalski’s Horse is likely genetically viable and therefore worthy of conservation.

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Bibliographic information: Ludovic Orlando et al. Recalibrating Equus evolution using the genome sequence of an early Middle Pleistocene horse. Nature, published online June 26, 2013; doi: 10.1038/nature12323