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CRISPR/Cas9 Used to Create Knockout Chickens

NEW YORK (GenomeWeb) – Scientists have used CRISPR/Cas9 to create genetically modified chickens for the first time, according to a new study published in Scientific Reports.

Led by Isao Oishi of Japan's National Institute of Advanced Industrial Science and Technology and Takahiro Tagami of the National Agriculture and Food Research Organization's Institute of Livestock and Grassland Science, the team reported it had to overcome difficulty in accessing the chicken zygotes in order to edit two genes for proteins found in egg whites, ovalbumin (OVA) and ovomucoid (OVM).

The scientists achieved editing efficiencies of greater than 90 percent for both targets in primordial germ cells (PGCs). They then implanted OVM knockout PGCs into embryos that developed into chimeric roosters. Subsequent genetic crosses led to ovomucoid homozygous mutant offspring.

"These results demonstrate that the CRISPR/Cas9 system is a simple and effective gene-targeting method in chickens," the authors wrote. Their results could have agricultural and industrial applications, particularly in reducing allergy concerns for people who might have an immune response to foods and vaccines containing egg whites.

Chickens, the authors said, have been particularly resistant to genome modification. Gene knockout in Gallus gallus domesticus has only been recently developed. In 2013, scientists from Crystal Bioscience reported successfully knocking out genes using sequence-specific nucleases and homologous recombination. And in 2014, Korean scientists used transcription activator-like effector nucleases (TALENs) to knock out OVA.

Creating double knockout birds that won't produce OVA or OVM is important because chicken eggs are used in many food products as well as in the manufacture of vaccines for viruses, notably influenza. While most people grow out of it by adulthood, egg allergies affect up to 3 percent of all young children by some estimates. The protein OVA is more abundant but OVM is the dominant allergen, the authors noted.

Double-knockout hens could produce hypoallergenic eggs, which might not affect individuals with egg-white allergies. Using gene editing to remove OVM might prove more effective than existing methods to reduce allergens in egg whites, since the OVM protein is stable and resistant to degradation by heat or proteolysis.

"It will be of immediate interest to test whether eggs from the homozygous knockout hens show reduced allergenicity," the authors said. They added that it might be possible to cross OVA and OVM mutant chickens to create birds lacking multiple allergy-related genes.