Biallelic modification of IL2RG leads to severe combined immunodeficiency in pigs.
BACKGROUND
Pigs with SCID can be a useful model in regenerative medicine, xenotransplantation, and cancer cell transplantation studies. Utilizing genome editing technologies such as CRISPR/Cas9 system allows us to generate genetically engineered pigs at a higher efficiency. In this study, we report generation and phenotypic characterization of IL2RG knockout female pigs produced through combination of CRISPR/Cas9 system and SCNT. As expected, pigs lacking IL2RG presented SCID phenotype.


METHODS
First, specific CRISPR/Cas9 systems targeting IL2RG were introduced into developing pig embryos then the embryos were transferred into surrogates. A total of six fetuses were obtained from the embryo transfer and fetal fibroblast cell lines were established. Then IL2RG knockout female cells carrying biallelic genetic modification were used as donor cells for SCNT, followed by embryo transfer.


RESULTS
Three live cloned female piglets carrying biallelic mutations in IL2RG were produced. All cloned piglets completely lacked thymus and they had a significantly reduced level of mature T, B and NK cells in their blood and spleen.


CONCLUSIONS
Here, we generated IL2RG knockout female pigs showing phenotypic characterization of SCID. This IL2RG knockout female pigs will be a promising model for biomedical and translational research.
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
Jung-Taek
JT
Kang
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
Bumrae
B
Cho
Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, USA.
Junghyun
J
Ryu
Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, USA.
Caitlin
C
Ray
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
Eun-Jin
EJ
Lee
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
Yun-Jin
YJ
Yun
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
SunMi
S
Ahn
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
JinSeok
J
Lee
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea.
Dal-Young
DY
Ji
Department of Large Animal Clinical Sciences, Virginia-Maryland College of Veterinary Medicine, Virginia Tech, Blacksburg, VA, USA.
Nathaniel
N
Jue
Department of Large Animal Clinical Sciences, Virginia-Maryland College of Veterinary Medicine, Virginia Tech, Blacksburg, VA, USA.
Sherrie
S
Clark-Deener
Department of Animal and Poultry Sciences, Virginia Tech, Blacksburg, VA, USA. kiholee@exchange.vt.edu.
Kiho
K
Lee
MGENPLUS Biotechnology Research Institute, Seoul, 08511, South Korea. parkmgen@hanmail.net.|Department of Animal Science & Technology, Sunchon National University, Suncheon, 57922, South Korea. parkmgen@hanmail.net.
Kwang-Wook
KW
Park
D000483:Alleles; D000818:Animals; D005260:Female; D055786:Gene Knockout Techniques; D005818:Genetic Engineering; D053631:Interleukin Receptor Common gamma Subunit / Q000235:genetics* / Q000502:physiology; D023421:Models, Animal; D016511:Severe Combined Immunodeficiency / Q000235:genetics / Q000662:veterinary*; D013552:Swine; D013553:Swine Diseases / Q000235:genetics*
D053631:Interleukin Receptor Common gamma Subunit
CRISPR/Cas9; IL2RG; Knockout pigs; Severe Combined Immunodeficiency (SCID); Somatic cell nuclear transfer (SCNT)