Replenish thylacine DNA


The thylacine (left) was Australia’s only marsupial predator to survive until European settlement. Its peculiar appearance has led to the collection of many specimens by scientists. One of them, C5757 (right), was used to sequence the thylacine genome. Credit: University of Melbourne

The DNA of extinct species opens a window on their biology, their evolution and their natural history. However, after an animal dies, its DNA invariably breaks into small pieces, making it difficult to reconstruct by computer.

Today, the new TIGRR lab, a research center focused on marsupial conservation and restoration, has used a clever approach to produce the most complete genome of the thylacine, also known as the Tasmanian tiger, to date. day.

This represents a significant step forward in our long-term goal of resurrecting the thylacine through a process called de-extinction.

Australia has one of the highest mammalian extinction rates in the world and the Tasmanian tiger is an emblem of this ongoing crisis.

The thylacine was once Australia’s largest marsupial predator, but was hunted to extinction in the early 20th century by settlers, enticed by a bounty of £1. Best known for its tiger stripes and dog-like head, the thylacine was captured in haunting photographs and newsreels before its extinction.

In 2018, our team, led by Professor Andrew Pask, published the first genome sequence of the thylacine.

To do this, we extracted DNA from a young pocket specimen at the Melbourne museum that had been stored in alcohol for over 100 years. We then used DNA sequencers to read the nearly three billion nucleotide “letters” of his genetic information and reassembled them computationally.

This genomic resource has allowed us and our collaborators to confirm the evolutionary relationships of thylacine with living marsupials, probe the genetic basis of its physical similarities to canids (dogs and their relatives), and show that thylacine had already suffered a long-term decline in membership. over several thousand years.

While the thylacine genome assembly project contained the overwhelming majority of its genetic information, we weren’t able to piece it all together.

In a living animal, DNA is organized into several chromosomes, each of which is a long DNA molecule made up of millions of letters. But due to thylacine DNA degradation over the past century, we’ve only been able to piece together chunks of several thousand letters at a time.

Today, advances in DNA assembly techniques and the explosion of high-quality reference genomes from living thylacine relatives have allowed us to construct a novel chromosomal-scale genome for thylacine .

In our study, published in the peer-reviewed journal Biology and evolution of the genomewe have combined two distinct approaches.

The first, called de novo assembly, uses complex algorithms to find overlapping stretches of DNA produced by a sequencing machine.

The second, called reference-guiding, uses the high-quality genomic sequence of a living parent as a template to help us assemble small fragments into full-length chromosomes.

In our case, we were able to align thylacine DNA sequences against the closely related Tasmanian Devil chromosomes with incredible efficiency and accuracy. This was possible due to a unique quirk in their biology.

Both the thylacine and the devil belong to the carnivorous marsupial order – Dasyuromorphia – a group that is notable for having some of the best conserved chromosomes among mammals. Therefore, our novel thylacine genome assembly is nearly of the same quality as that of recently sequenced marsupial species such as the fat-tailed dunnart, tammar wallaby, and ground couscous.

A high quality genome is essential for thylacine de-extinction. This is because there are no living thylacine cell lines, so we can’t just clone a thylacine, like Dolly the sheep.

Instead, we will need to use genome editing with techniques like CRISPR-Cas9 to reconstruct the thylacine genome in marsupial stem cells. Our new genome assembly will help the TIGRR lab team map around 90% of these genome edits, and further improvements will take us the rest of the way.

Our new genome assembly also allows us to dig deeper into the genetic diversity of the thylacine before its extinction. For example, we have now shown that the thylacine has the lowest diversity of any marsupial species we have examined.

This discovery confirms a previous study to us that used mitochondrial DNA, but at the genome scale, offering greater power and precision. Additionally, this is consistent with our 2018 demographic analysis, which showed that the thylacine had suffered a long-term decline in numbers, likely representing a genetic bottleneck.

Low genetic diversity can have significant impacts on a species’ fitness.

Inbreeding can cause harmful recessive disorders to manifest in a population. Additionally, low diversity can make species vulnerable to disease and unable to adapt to a changing environment.

However, although low diversity and genetic health are intimately linked, they are not synonymous. Some species that have suffered deep bottlenecks have been brought back from the brink.

The Arabian oryx (Oryx leucoryx) became extinct in the wild in the 1970s and was kept for some time only in captivity, with a population of only nine individuals. From this tiny cohort, the species grew to over 6,000 individuals, with about 1,000 in the wild.

Today, the Arabian oryx is no longer considered an endangered species.

Going forward, we aim to continue to improve our thylacine genome at the chromosomal level and use this resource to better understand the relationship between low thylacine diversity and its genetic health. This can be done by sequencing the DNA of hundreds of thylacines kept in museums around the world.

Studies like this are critical to TIGRR Lab’s long-term goal of resurrecting Australia’s vital apex predator through genome engineering while ensuring the health and viability of their future populations.

The nine steps to bringing Australia’s extinct thylacine back

More information:
Charles Feigin et al, A chromosome-scale hybrid genome assembly of the extinct Tasmanian tiger (Thylacinus Cynocephalus), Biology and evolution of the genome (2022). DOI: 10.1093/gbe/evac048

Provided by the University of Melbourne

Quote: Thylacine DNA reconstruction (2022, March 31) Retrieved March 31, 2022 from

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