Over the course of two decades, Japanese researchers repeatedly cloned mice, but the experiment revealed the limits of this approach: by the 58th “iteration,” they could no longer obtain viable offspring. As reported in a paper published in Nature Communications, the key reason was not disrupted epigenetic regulation but the accumulation of genetic mutations.

The results are directly tied to a fundamental question in biology: why living organisms need sexual reproduction. It is believed that without gene recombination, the so-called Muller’s ratchet is set in motion—the gradual buildup of harmful mutations that over time reduces a population’s viability. Although nature includes organisms capable of asexual reproduction, mammals are fully dependent on sexual reproduction. Cloning made it possible to test what happens if one tries to bypass this mechanism.

How the experiment was conducted

The work by Sayaka Wakayama and Teruhiko Wakayama’s group (University of Yamanashi) began in 2005. The scientists transferred the nucleus of a mouse somatic cell into an egg cell, raised the clone, and then repeated the procedure using cells from the animal that had been produced. The early generations reproduced relatively successfully, but after the 27th generation fertility began to decline. Ultimately, the 58th generation was nonviable: the newborn mice did not survive even a day.

Why it wasn’t epigenetics

Comparative analysis showed that the clones did indeed differ from ordinary mice (for example, larger placentas and higher body mass), but these changes did not increase over time. Epigenetic markers and the animals’ overall condition remained stable, and the surviving individuals lived normally. This led the authors to conclude that the reason the clone series stopped was not epigenetic failures.

What tests via sexual reproduction showed

Additional experiments found that even with normal fertilization, eggs from cloned mice showed reduced quality: most embryos failed to reach the blastocyst stage. However, a small fraction still developed, suggesting the possibility of a partial “reset” of accumulated problems when returning to sexual reproduction.

Mutation accumulation as the key limiting factor

Genetic analysis confirmed that the number of mutations increased with each cloning cycle, and after the 25th generation major chromosomal abnormalities also began to appear. This is explained by the fact that cloning used somatic cells that had already accumulated changes, and the absence of recombination did not allow the genome to be effectively “cleansed” of harmful mutations.

At the same time, the surviving animals did not show a noticeable decline in health or a shortened lifespan. This is likely due to strong selection at early developmental stages: only those embryos whose mutation burden remained compatible with life survived.

Conclusions and practical significance

The study provided a clear demonstration of Muller’s ratchet in mammals. The authors emphasize that the main problem with long-term cloning is linked not so much to technical or epigenetic difficulties as to the lack of genetic “renewal” mechanisms characteristic of sexual reproduction.

Nevertheless, cloning is still considered a promising tool—for example, for conserving rare animal species—despite the limitations identified.