• ISSN 1674-8301
  • CN 32-1810/R
Volume 31 Issue 5
Sep.  2017
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Article Contents
Ze Li, Hai-Yuan Yang, Ying Wang, Man-Ling Zhang, Xiao-Rui Liu, Qiang Xiong, Li-Ning Zhang, Yong Jin, Li-Sha Mou, Yan Liu, Rong-Feng Li, Yi Rao, Yi-Fan Dai. Generation of tryptophan hydroxylase 2 gene knockout pigs byCRISPR/Cas9-mediated gene targeting[J]. The Journal of Biomedical Research, 2017, 31(5): 445-452. doi: 10.7555/JBR.31.20170026
Citation: Ze Li, Hai-Yuan Yang, Ying Wang, Man-Ling Zhang, Xiao-Rui Liu, Qiang Xiong, Li-Ning Zhang, Yong Jin, Li-Sha Mou, Yan Liu, Rong-Feng Li, Yi Rao, Yi-Fan Dai. Generation of tryptophan hydroxylase 2 gene knockout pigs by CRISPR/Cas9-mediated gene targeting[J]. The Journal of Biomedical Research, 2017, 31(5): 445-452. doi: 10.7555/JBR.31.20170026

Generation of tryptophan hydroxylase 2 gene knockout pigs by CRISPR/Cas9-mediated gene targeting

doi: 10.7555/JBR.31.20170026
Funds:

This work was supported by a grant from the National Natural Science Foundation of China (No. 81570402), a grant from the Jiangsu Key Laboratory of Xenotransplantation (BM2012116), and grants from the Sanming Project of Medicine in Shenzhen, the Fund for High Level Medical Discipline Construction of Shenzhen (No. 2016031638), and the Shenzhen Foundation of Science and Technology (No. JCYJ20160229204849- 975 and GCZX2015043017281705), and grant from the National Basic Research Program of China (2015CB55- 9200).

  • Received: 2017-03-05
  • Revised: 2017-03-29
  • Issue Date: 2017-09-22
  • Unbalanced brain serotonin (5-HT) levels have implications in various behavioral abnormalities and neuropsychiatric disorders. The biosynthesis of neuronal 5-HT is regulated by the rate-limiting enzyme, tryptophan hydroxylase-2 (TPH2). In the present study, the clustered regularly interspaced short palindromic repeat (CRISPR)/ CRISPR-associated (Cas) system was used to target the Tph2 gene in Bama mini pig fetal fibroblasts. It was found that CRISPR/Cas9 targeting efficiency could be as high as 61.5%, and the biallelic mutation efficiency reached at 38.5%. The biallelic modified colonies were used as donors for somatic cell nuclear transfer (SCNT) and 10 Tph2 targeted piglets were successfully generated. These Tph2 KO piglets were viable and appeared normal at the birth. However, their central 5-HT levels were dramatically reduced, and their survival and growth rates were impaired before weaning. These Tph2 KO pigs are valuable large-animal models for studies of 5-HT deficiency induced behavior abnomality.

     

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