An albino opossum proves that CRISPR works in marsupials too

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MIT Technology Review

While kangaroos and koalas are better known, researchers studying marsupials often use possums in laboratory experiments because they are smaller and easier to care for. Gray short-tailed opossums, the species used in the study, are related to the white-faced North American opossums, but they are smaller and have no pouch.

Riken’s researchers used CRISPR to delete or turn off a gene that codes for pigment production. Targeting this gene meant that if the experiments worked, the results could be seen at a glance: the opossums would be albino if both copies of the gene were turned off, and speckled or mosaic if a single copy was removed.

The resulting litter included an albino opossum and a mosaic opossum (pictured above). The researchers also bred the two, resulting in a litter of fully albino possums, showing that the coloring was an inherited genetic trait.

The researchers had to overcome some hurdles in order to edit the opossum genome. First they had to find out when the hormone injections would take them to prepare the animals for pregnancy. The other challenge was that shortly after fertilization, marsupials developed a thick layer around them called the mucous membrane. This makes it difficult to inject the CRISPR treatment into the cells. On her first attempts, needles either wouldn’t penetrate the cells or would damage them so the embryos couldn’t survive, Kiyonari says.

The researchers realized that it would be much easier to inject earlier, before the coating around the egg got too hard. By changing when the lights were turned off in the labs, the researchers got the possums to mate later in the evening so the eggs would be ready to work in the morning, about a day and a half later.

The researchers then used a tool called a piezoelectric drill that uses electrical charge to make it easier to penetrate the membrane. This helped them inject the cells without damaging them.

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

Opossums have been used as laboratory animals since the 1970s, and researchers have been trying to modify their genes for at least 25 years, says VandeBerg, who began attempting the first laboratory opossum colony in 1978. They were also the first marsupials to have their genome 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 marsupial genomes that we don’t have, which puzzles them,” says Rob Miller, an immunologist at the University of New Mexico who uses possums in his research.

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