At the heart of Colossal Biosciences’ groundbreaking de-extinction of the dire wolf lies a sophisticated understanding of genetic architecture – the complex relationship between genes and physical traits. By identifying and editing 14 crucial genes, Colossal’s scientists were able to transform gray wolf cells into dire wolf proxies that express key characteristics of their extinct counterparts. This precise genetic engineering, involving 20 distinct edits across these 14 genes, represents a remarkable feat of genomic precision and offers fascinating insights into the molecular basis of species traits.
The Genetic Blueprint
Colossal’s approach began with extracting and analyzing ancient DNA from dire wolf fossils – a 13,000-year-old tooth from Sheridan Pit, Ohio and a 72,000-year-old inner ear bone from American Falls, Idaho. Using advanced computational methods, the team assembled what they describe as “high quality ancient genomes” with much greater coverage than previously achieved.
This genetic information allowed them to identify the specific genes and variants that made dire wolves unique among canids. According to Colossal’s documentation, “We identified gene variants specific to dire wolves” and “determined that dire wolves had a white coat color, and long thick fur” – traits that couldn’t be determined from fossils alone.
With this genetic blueprint in hand, the team selected 14 genes to target for editing, focusing on those that would resurrect key dire wolf traits while ensuring the health and viability of the resulting animals.
Coat Color and Texture Genes
Several of the targeted genes govern coat characteristics – a defining feature of dire wolves that differentiates them from their gray wolf relatives.
CORIN: The White Coat Gene
One of the most visually striking genes edited was CORIN, a serine protease expressed in hair follicles that influences coat color through its effects on the agouti pathway. According to Colossal’s research, “The dire wolf CORIN variants impact pigmentation in a way that leads to a light coat color.”
This gene helps explain why the de-extinct dire wolves have their distinctive white coats – a trait that may have evolved in response to Ice Age conditions. The snowy white appearance of the pups Romulus, Remus, and Khaleesi directly reflects the successful editing of this key pigmentation gene.
Pigmentation Gene Adaptations
Interestingly, Colossal’s approach to coat color genes demonstrates their careful balancing of genetic authenticity with animal welfare. Their analysis identified dire wolf-specific variants in three essential pigmentation genes: OCA2, SLC45A2, and MITF, which “directly impact the function and development of melanocytes.”
However, the team discovered that these specific variants could potentially cause health problems in gray wolves. “While these variants would have led to a light coat in dire wolves, variation in these genes in gray wolves can lead to deafness and blindness,” explains their documentation.
Rather than risking these health issues, Colossal chose an alternative genetic pathway to achieve the same coat color safely: “The team therefore engineered a light colored coat in Colossal’s dire wolves via a path known to be safe in gray wolves: by inducing loss-of-function to MC1R and MFSD12.” These genes influence the expression of eumelanin (black) and pheomelanin (red) pigments, achieving the lighter coat color without health risks.
Dr. Elinor Karlsson, Associate Professor at UMass Chan Medical School, praised this approach: “By choosing to engineer in variants that have already passed evolution’s clinical trial, Colossal is demonstrating their dedication to an ethical approach to de-extinction.”
Size and Skeletal Structure Genes
Another set of targeted genes governs the dire wolf’s larger size and distinctive skeletal structure – traits that made them formidable predators in the Ice Age.
HMGA2: The Body Size Gene
HMGA2 is a gene “directly associated with body size in dogs and wolves” according to Colossal’s research. This gene is part of a multi-gene regulatory module that influences size as well as ear, skull, and facial morphology.
By editing dire wolf-specific variants into this gene, Colossal helped ensure that their de-extinct dire wolves would develop the larger, more robust build characteristic of the species. At just six months old, the male pups already weigh approximately 80 pounds – showcasing the effectiveness of these size-related edits.
MSRB3: Shaping Skull and Ears
Another gene in the same regulatory module, MSRB3, “has been linked to variation in ear and skull shape among canines and other mammals.” Dire wolves had distinctively broad heads and robust skulls adapted for powerful biting forces – traits that appear to be developing in the de-extinct pups thanks to the successful editing of this gene.
LCORL: The Growth Regulator
LCORL is a transcription factor that regulates gene expression and has been linked to body size variation in many species. According to Colossal’s documentation, “The dire wolf has three changes to the LCORL protein sequence that are predicted via 3D modeling to alter the way the protein folds precisely at the location where LCORL should bind to a major gene silencing complex known as the PRC2 domain.”
Interestingly, large dog breeds already have a variant of LCORL that is missing this PRC2 domain. Given the similarity in phenotypic impact between the dire wolf variant and the one found in large dog breeds, Colossal opted to use “the protein that is found in the largest grey wolves” for this edit, balancing the desired size effect with genetic compatibility.
Muscular Development Genes
Dire wolves were not just larger than gray wolves but also more muscular, with adaptations for taking down large Ice Age prey like horses and bison.
Colossal’s documentation mentions that they identified “multiple genes undergoing positive selection that are linked to dire wolf skeletal, muscular, circulatory, and sensory adaptation.” While specific muscular development genes aren’t named in the available materials, the reference to genes linked to “muscular adaptation” indicates that some of the 14 targeted genes influence muscle mass and composition.
The early development of the dire wolf pups suggests these edits are having their intended effect, with observers noting their “hefty builds” even at a young age. As the wolves mature, the full impact of these muscular development gene edits will likely become more apparent.
Vocalization and Behavioral Genes
Perhaps most fascinating are the genes that influence behavioral traits, including the dire wolf’s unique vocalization patterns.
According to TIME Magazine’s reporting, Colossal identified genes influencing “the dire wolf’s larger size, more muscular build, wider skull, bigger teeth, thick light-colored coat, and even its unique howling vocalizations.”
This suggests that some of the 14 targeted genes affect neural development and vocalization patterns, potentially giving the de-extinct dire wolves distinctive howls that differ from those of modern gray wolves. Such behavioral traits are rarely preserved in the fossil record, making these genetic insights particularly valuable for understanding extinct species’ behavior.
Regulatory Region Edits
Beyond editing protein-coding genes, Colossal also made changes to regulatory regions of the genome – stretches of DNA that control when and how genes are expressed rather than coding for proteins themselves.
Their documentation notes that they identified “dire wolf specific variants in regulatory regions that alter the expression of genes.” These regulatory edits are crucial for recreating complex dire wolf traits that emerge from the coordinated expression of multiple genes.
Specifically, they edited “dire wolf-specific variants in a multi-gene regulatory module that has been linked to variation in body size as well as ear, skull, and facial morphology. The region encodes eight genes that establish species-specific constraints in skeletal size and structure.”
This sophisticated approach to genetic engineering acknowledges that many species-defining traits don’t arise from single genes but from complex regulatory networks that orchestrate development.
A New Benchmark in Precision Editing
Together, these 14 genes and the 20 edits made across them represent an unprecedented achievement in multiplex genetic engineering. According to Colossal, this is “the largest number of precise genomic edits in a healthy vertebrate so far” – surpassing their previous record of 8 edits in their “woolly mouse” demonstration from early 2025.
What makes this achievement particularly remarkable is that 15 of the 20 edits involved “ancient gene variants that have not existed in over 12,000 years” – bringing back genetic sequences that disappeared from Earth with the extinction of the dire wolf.
The successful birth and healthy development of the dire wolf pups demonstrates that these complex genetic modifications are not just theoretically possible but practically achievable. As Harvard geneticist and Colossal co-founder Dr. George Church noted, this represents “a capability that is growing exponentially.”
Implications Beyond Dire Wolves
The identification and successful editing of these 14 key genes has implications that extend far beyond the dire wolf itself. By demonstrating the ability to precisely modify multiple genes to resurrect extinct traits, Colossal has established a proof of concept for more ambitious de-extinction efforts, including their planned resurrection of the woolly mammoth by 2028.
Moreover, the approaches developed for identifying and editing these dire wolf genes can be applied to genetic rescue efforts for endangered species. The same technologies used to resurrect dire wolf traits could potentially be used to restore genetic diversity in inbred populations or introduce adaptive traits to help species cope with environmental changes.
As we continue to observe the growth and development of Romulus, Remus, and Khaleesi, these 14 carefully selected genes will express themselves more fully, bringing the distinctive characteristics of the dire wolf back into the living world after an absence of thousands of years – a testament to both the power of modern genetic engineering and the remarkable conservation of genetic information across evolutionary time.
