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| Establishment of RPE65 Gene-edited Dog (Canis lupus familiaris) Using TALEN Technology |
| XU Zheng1,2, LEE Chen-Cheng2, LAI Wei-Chung2, XU Bo2, QIAN Yu-Yu2, CHEN Chen2, WANG Jing2, ZHANG Yong2, FENG Chong2,*, WEI Yan-Ming1,* |
1 College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China;
2 Gansu Key Laboratory of Animal Generational Physiology and Reproductive Regulation, Lanzhou 730070, China;
3 Aurora Bioscience Co., Ltd, Suzhou 215400, China |
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Abstract The retinal pigment epithelium 65 (RPE65) gene plays an important role in the visual cycle; when the RPE65 gene is mutated, there is a loss of cone and rod cell function, and loss of vision in severe patients. In order to construct an animal disease model with RPE65 gene mutation; this study utilized transcription activator-like effector nucleases (TALEN) gene editing technology to construct expression vectors for Exon3 and Exon7 of the RPE65 gene in Beagle dogs (Canis lupus familiaris), and combined with microscopic manipulation to create RPE65 gene-edited dogs. The dogs' visual function was tested by clinical observation, routine ophthalmologic examination, fundus photography (FP), optical coherence tomography (OCT), and full-field electroretinography (ERG). T7E1 digestion and gene sequencing results showed that 2 RPE65 gene-edited male dogs were finally obtained. Ophthalmic phenotypic analysis showed that the fundus blood vessels of RPE65 gene-edited dogs were sparse and slender. The ERG results showed that the amplitude of the a-wave and b-wave potentials in photoreceptor cells decreased, and the peak time prolonged, indicating impaired retinal function. In this study, RPE65 gene-edited dogs were constructed by using TALEN gene editing technology with visual dysfunction, which may provide a reference for studies related to retinal diseases induced by mutations in the RPE65 gene, and also provide a good animal disease model for the development of therapeutic treatments and the screening of clinical drugs.
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Received: 06 April 2023
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Corresponding Authors:
* Corresponding authors, weiym@gsau.edu.cn; fengchong@joinn-lab.com
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