In a preserve at an undisclosed location, two so-called dire wolves roam, eat, and play, blissfully unaware that their existence has ignited a heated scientific and ethical debate. According to Colossal Biosciences, a company focused on de-extinction projects, who created the now six-month-old animals, this is the first time a species has been brought back from extinction.

Romulus and Remus are genetically identical but were birthed by different mothers. To accommodate their expected size, Colossal chose large mixed-breed dogs to carry the pups.
Colossal Biosciences
Named after the Romulus and Remus of Roman mythology—twin human brothers raised by a she-wolf—the Aenocyon dirus pups were created using multiplexed CRISPR editing of the genome of the extant gray wolf, Canis lupus. While some researchers hail this development as a breakthrough in conservation science, many are hesitant to jump on the de-extinction bandwagon. Others suggest that the animals are not dire wolves at all, but rather genetically modified gray wolves or hybrids.
So, what, exactly, does it mean to be a dire wolf? Colossal recently sequenced the first complete genome of the dire wolf from two fossils: a 13,000-year-old tooth and a 72,000-year-old inner ear bone. The currently unpublished analysis, which built on chief science officer Beth Shapiro’s 2021 study in Nature, revealed that the dire wolf genome shares 99.5 percent similarity with that of the gray wolf.1
Using endothelial progenitor cells taken from the blood of a gray wolf, the Colossal team incorporated 20 edits across 14 genes to produce traits that differentiate the dire wolf from the gray wolf. They then removed the nucleus from the edited cell and inserted it into the denucleated egg cell of a domestic dog. After the resulting cell developed into an embryo, it was implanted into a large domestic dog, who acted as a surrogate.
The genes Colossal edited were chosen because they influenced phenotypic factors, including size, skull shape, and musculature, as well as hair color, texture, and length. Herein lies the wolves’ perceived identity crisis. Paleoecologist Jacquelyn Gill of the University of Maine, who was not involved in the study, told Scientific American that the animals are not true dire wolves: “I have more than 14 Neandertal genes in me, and we wouldn’t call me a Neandertal.”

Colossal’s de-extinction program was based on editing the genome of the gray wolf to incorporate regions of DNA from the dire wolf that are responsible for several of its characteristic features, such as its large size.
Colossal Biosciences
Shapiro and her colleagues disagree. “If we can look at this animal and see what it’s doing, and it looks like a dire wolf and acts like a dire wolf, I’m going to call it a dire wolf,” Shapiro said in an interview with Wired.
Long-term plans for Romulus and Remus, and their younger sister Khaleesi, haven’t been decided. Colossal says they are attempting to create more of the animals but don’t have plans for a breeding program. Scientists like Gill are skeptical of any potential introduction of dire wolves into the wild.
“I think the dire wolf is absolutely a symbol of hope,” bioethicist and Colossal advisor Alta Charo said in a video released by Colossal, noting that the technologies developed for the dire wolf de-extinction program have yielded a range of benefits that apply to conservation. Beyond the de-extinction of the dire wolf, Colossal hopes to inform conservation efforts for living species that are currently endangered, like the red wolf Canis rufus. Critically endangered, the red wolf has a paucity of genetic variation that Colossal intend to supplement with four pups they have produced using their cloning methods.
- Perri AR, et al. Dire wolves were the last of an ancient New World canid lineage. Nature. 2021;591(7848):87-91.