It was in June 2019 that the birth of a specific batch of gene-edited piglets was announced to the world. It was a significant step in the quest to achieve commercial production of male pigs that never reach adolescence, and therefore do not need castration to prevent boar taint.
Created through an alliance between the Acceligen unit of Recombinetics and Hendrix Genetics, the milestone built on previous research that was funded partly by a 2017 grant from the Foundation for Food and Agriculture Research to Recombinetics, allotted for the purposes of developing a way to end physical castration.
Gene edit details
This particular gene edit relates to stopping expression of a gene that triggers the release of hormones necessary for sexual maturation. The gene, called KISS1, is expressed in the pig's pituitary gland during puberty, and triggers certain cells to mature and begin producing hormones, which propel puberty forward as the animal grows older.
The genetic basis for the hormones that relate to growth and maturation in humans has already been studied extensively, notes Tad Sonstegard, who leads the piglet research and is chief executive and scientific officer at Acceligen, Recombinetics' agriculture division.
"Livestock scientists have also studied the KISS1 gene in pigs for quite some time, to investigate how puberty in pigs might be accelerated, to increase fertility and to determine the effects on sow longevity," Sonstegard says.
Research conducted by Björn Peterson of the Friedrich Loeffler Institute in Greifswald, Germany, and his colleagues published last year investigated how stopping the expression of pig genes on the Y chromosome can produce males that develop with female attributes — and perhaps no boar taint.
In the Recombinetics-Acceligen and Hendrix Genetics alliance research, the gene edits were completed in pig embryos at the single-cell stage, harvested from slaughtered sows.
A Cas9 protein is introduced into the cell, which seeks out the gene-editing tool CRISPR "guide RNA" that marks the portion of the embryo's genome where KISS1 is encoded. This complex then cuts the gene. Afterward, the embryo's DNA repair mechanisms step in and fix this gap.
The embryos are then placed into recipient gilts. When the piglets are born, a blood or tissue sample is taken, and through the well-established polymerase chain reaction method, the KISS1 segment of DNA targeted by CRISPR is sequenced to confirm the edit.
To achieve commercial production of male pigs that do not reach adolescence (and therefore do not have boar taint), male pigs obviously need to receive the gene edit from their parents. However, because the parent nucleus stock would also have this gene edit and therefore would stay preadolescent themselves, an effective and efficient way must be found to make both male and female parent stock fertile.
The strategy that will be used here involves supplying an analog of KISS1 to the pigs. This should cause maturation of the gonads and subsequent production of the desired germ cells (eggs and sperm). "We will be doing a number of measurements related to growth and gonad development," Sonstegard explains. "Meanwhile, we will also test these animals, about 30 of them, for boar taint components; there should be none."
In this experimental setting, the KISS1 will be delivered by experimental protocols approved by animal ethics review. "If the first few experimental tries are promising, then we would work to robustly deliver the analog in a commercial nucleus farm setting, using a method that's both efficient and economical," Sonstegard says.
Sonstegard and his colleagues also need to determine whether deletion of both copies of the KISS1 gene is required for pigs to stay preadolescent. "Some pigs we breed from our founding population of edited animals would have one functional KISS1 gene," he says.
"We will attempt to identify if pigs that carry only one functional copy of the gene still express puberty normally, or is puberty delayed and/or is fertility reduced due to the potential reduced expression of KISS1? Our work should shed some light on different alternatives for pig breeding, as well as present new information on the reproductive biology of both female and male pigs optimally bred for commercial pork production."
After that, the team will move on to study how preadolescent males and females perform in comparison to conventional pigs: that is, their commercial viability.
"In order for widespread commercial production, traits like feed efficiency, sow performance and meat quality cannot be compromised," Sonstegard says.