Generation of TALEN-mediated FH knockout rat model
Dandan YuYali ZhongXiaoran LiYaqing LiXiaoli LiJing CaoZhirui FanHuijie FanLong YuanBenling XuYuan YuanHongquan ZhangZhenyu JiJianguo WenMingzhi ZhangJahn M. NeslandZhenhe Suo
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// Dandan Yu 1, 9, 10 , Yali Zhong 1 , Xiaoran Li 9, 10 , Yaqing Li 1 , Xiaoli Li 1 , Jing Cao 1, 2 , Zhirui Fan 1 , Huijie Fan 1 , Long Yuan 3 , Benling Xu 4 , Yuan Yuan 5 , Hongquan Zhang 6 , Zhenyu Ji 7 , Jian-Guo Wen 8 , Mingzhi Zhang 1 , Jahn M. Nesland 9, 10 , Zhenhe Suo 1, 9, 10 1 Department of Oncology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China 2 Departments of Pathology, The Third Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan, China 3 Department of Surgery, The Affiliated Cancer Hospital of Zhengzhou University, Zhengzhou, Henan, China 4 Department of Cancer Biotherapy, The Affiliated Cancer Hospital of Zhengzhou University, Zhengzhou, Henan, China 5 Department of Pathology, Capital Medical University, Beijing, Fengtai, China 6 Department of Anatomy, Histology and Embryology, Peking University Health Science Center, Beijing, Fengtai, China 7 Department of Oncology, Henan Academy of Medical and Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, Henan, China 8 Institute of Clinical Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou University, Zhengzhou, Henan, China 9 Department of Pathology, The Norwegian Radium Hospital, Oslo University Hospital, Oslo, Norway 10 Department of Pathology, Institute of Clinical Medicine, Faculty of Medicine, University of Oslo, Oslo, Norway Correspondence to: Zhenhe Suo, email: zhenhes@medisin.uio.no Keywords: TALEN, fumarate hydratase, gene technology, obesity, heterozygote Received: February 23, 2016 Accepted: August 09, 2016 Published: August 19, 2016 ABSTRACT Transcription activator-like effector nucleases (TALENs) are valuable tools for precise genome engineering of laboratory animals. Here we utilized this technique for efficient site-specific gene modification to create a fumarate hydratase (FH) gene knockout rat model, in which there was an 11 base-pair deletion in the first exon of the FH gene in 111 rats. 18 live-born targeted mutation offsprings were produced from 80 injected zygotes with 22.5% efficiency, indicating high TALEN knockout success in rat zygots. Only heterozygous deletion was observed in the offsprings. Sixteen pairs of heterozygous FH knockout (FH+/–) rats were arranged for mating experiments for six months without any homozygous KO rat identified. Sequencing from the pregnant rats embryo samples showed no homozygous FH KO, indicating that homozygous FH KO is embryonically lethal. Comparatively, the litter size was decreased in both male and female FH+/– KO rats. There was no behaviour difference between the FH+/– KO and the control rats except that the FH+/– KO male rats showed significantly higher body weight in the 16-week observation period. Clinical haematology and biochemical examinations showed hematopoietic and kidney dysfunction in the FH+/– KO rats. Small foci of anaplastic lesions of tubular epithelial cells around glomeruli were identified in the FH+/– kidney, and these anaplastic cells were comparatively positive for Ki67, p53 and Sox9, and such findings are most probably related to the kidney dysfunction reflected by the biochemical examinations of the rats. In conclusion, we have successfully established an FH+/– KO rat model, which will be useful for further functional FH studies.In Part I, this CRS report discusses the policy on “one China” since the United States began in 1971 to reach understandings with the People’s Republic of China (PRC) government in Beijing. Part II documents the evolution of the “one China” principle as articulated in key statements by Washington, Beijing, and Taipei. The U.S. policy on “one China” has evolved to cover three issues: sovereignty, peaceful resolution, and cross-strait dialogue.
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