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Sci Rep. 2018 May 16;8(1):7661. doi: 10.1038/s41598-018-25654-8.

Cattle with a precise, zygote-mediated deletion safely eliminate the major milk allergen beta-lactoglobulin.

Author information

1
AgResearch, Ruakura Research Centre, Hamilton, 3240, New Zealand.
2
Rowett Institute, Aberdeen, AB25 2ZD, United Kingdom.
3
MS3 Solutions Ltd., Ruakura Research Centre, Hamilton, 3240, New Zealand.
4
Recombinetics, St. Paul, MN, United States.
5
AgResearch, Ruakura Research Centre, Hamilton, 3240, New Zealand. goetz.laible@agresearch.co.nz.

Abstract

We applied precise  zygote-mediated genome editing to eliminate beta-lactoglobulin (BLG), a major allergen in cows' milk. To efficiently generate LGB knockout cows, biopsied embryos were screened to transfer only appropriately modified embryos. Transfer of 13 pre-selected embryos into surrogate cows resulted in the birth of three calves, one dying shortly after birth. Deep sequencing results confirmed conversion of the genotype from wild type to the edited nine bp deletion by more than 97% in the two male calves. The third calf, a healthy female, had in addition to the expected nine bp deletion (81%), alleles with an in frame 21 bp deletion (<17%) at the target site. While her milk was free of any mature BLG, we detected low levels of a BLG variant derived from the minor deletion allele. This confirmed that the nine bp deletion genotype completely knocks out production of BLG. In addition, we showed that the LGB knockout animals are free of any TALEN-mediated off-target mutations or vector integration events using an unbiased whole genome analysis. Our study demonstrates the feasibility of generating precisely biallelically edited cattle by zygote-mediated editing for the safe production of hypoallergenic milk.

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