Researchers from the Francis Crick Institute and the University of Kent used gene-editing technologies to create male-only and female-only mice litters, according to a study published in Nature Communications (1). The authors also suggested ways in which this method could be used to improve animal welfare in scientific research and agriculture.
There are many situations in research and agriculture where it would be desirable to have just females or just males. For example, reproductive studies require only animals of the gender being studied, while in farming, egg and milk production needs only female animals. Sadly, in many cases, the unwanted animals end up being culled.
“This work could have an immediate and valuable impact in scientific laboratories, as we’ve shown how it is safe and effective in mice, a common mammal used in medical and scientific research. While a lot of research needs both sexes, there are areas of study where only one is needed. For example, when studying the reproductive system, sex-specific diseases, or certain hormones”, said James Turner, group leader of the Sex Chromosome Biology Laboratory at the Crick says:
Using gene-editing tools, the team managed to find a way to inactivate embryos shortly after fertilisation, allowing only the desired sex to develop. This selection relies on the two elements of the CRISPR-cas9 system: the Cas9 enzyme that cuts the DNA in specific regions and the guide RNA that transports the Cas9 enzyme to the correct location.
The researchers added one of the elements to the father’s X or Y chromosome, which meant that it could only be inherited by males or females, respectively. The second element was added to the mother, and therefore inherited by all embryos. They decided to target a gene called Top1, which is essential for DNA repair. Embryos containing both parts of the CRISPR-cas9 system couldn’t develop beyond the 16 to 32 cells, whereas embryos with only one part developed normally.
The results were 100% effective, and litters were either all male or all female. To produce a male-only litter, the team edited the father’s X chromosome, and for a female-only, they edited the father’s Y chromosome.
“This method works as we split the genome editing process in half, between a male and female, and it is only when the two halves meet in an embryo through breeding that it is activated. Embryos with both halves cannot develop beyond very early cell stages”, said Charlotte Douglas from the Crick Institute. “We’ve also shown this process works successfully in different combinations – introducing either the Cas9 or the guide RNA elements onto the mother’s or father’s chromosomes.”
Surprisingly, the method did not actually result in litters that were 50% smaller due to the loss of one gender. The researchers believe this is because female mice produce more eggs than required, allowing some of them to be lost during early development. This means that in situations where only one sex is needed, fewer breeding animals will be required to produce the same number of the desired sex.
As the surviving offspring only contain one half of the CRISPR-cas9 system, this sex selection cannot be passed on to the next generation unless they are bred with an animal of the opposite sex that contains the other half. This is not the same as genetic engineering using gene drive methods, which were developed to spread a genetic mutation through the population.
“The implications of this work are potentially far-reaching when it comes to improving animal welfare, but should be considered at ethical and regulatory levels,” concluded Peter Ellis from the University of Kent. “In particular, before any potential use in agriculture, there would need to be extensive public conversation and debate, as well as changes to legislation. On the scientific side, there is also much work to be done over a number of years. Further research is needed, first to develop the particular gene-editing toolkits for different species, and then to check they are safe and effective.”
(1) Douglas, C. et al. (2021). CRISPR-Cas9 effectors facilitate generation of single-sex litters and sex-specific phenotypes. Nature Communications. 10.1038/s41467-021-27227-2.