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Although kangaroos and koalas are better known, marsupial researchers often use possums in laboratory experiments because they are smaller and easier to care for. The gray short-tailed possums, the species used in the study, are related to the white-faced North American possums, but they are smaller in size and do not have a pouch.

Researchers at Riken have used CRISPR to remove or knock out a gene that codes for pigment production. Targeting this gene meant that if the experiments worked, the results would be obvious at a glance: possums would be albinos if both copies of the gene were knocked out, and spotted or mosaic if one copy was deleted.

The result was a litter of one albino possum and one mosaic possum (pictured above). The researchers also crossed the two, resulting in a litter of albino possums, showing that the coloration is a hereditary genetic trait.

Researchers had to overcome several obstacles in order to edit the possum genome. First, they had to determine the timing of hormone administration in order to prepare the animals for pregnancy. Another problem was that soon after fertilization, marsupial eggs developed a thick layer around them called mucous membranes. This makes it difficult to introduce CRISPR processing into cells. According to Kiyonari, in their first attempts, the needles either failed to penetrate the cells or damaged them, so the embryos could not survive.

The researchers realized that it would be much easier to inject at an earlier stage, before the coating around the egg gets too stiff. By varying the times when the lights were turned off in the laboratories, the researchers forced the possums to mate later in the evening so that the eggs would be ready for work in the morning, about a day and a half later.

The researchers then used a tool called a piezoelectric drill, which uses an electrical charge to penetrate the membrane more easily. This helped them enter cells without damaging them.

“I think this is an incredible result,” says Richard Behringer, a geneticist at the University of Texas. “They have shown that it is possible. Now it’s time to get into biology, ”he adds.

“Opossums have been used as laboratory animals since the 1970s, and researchers have been trying to edit their genes for at least 25 years,” says Wandeberg, who began his attempts to create the first laboratory colony of possums in 1978. the genome was fully sequenced in 2007.

Comparative biologists hope that the ability to genetically modify possums will help them learn more about some of the unique aspects of marsupial biology that have yet to be deciphered. “We find genes and genomes from marsupials that we don’t have, so this creates some mystery as to what they’re doing,” says Rob Miller, an immunologist at the University of New Mexico who uses possums in his research. …