臺灣大學: 漁業科學研究所周宏農; 張繼堯莊韻蓓Chung, Yun-PeiYun-PeiChung2013-03-272018-07-062013-03-272018-07-062010http://ntur.lib.ntu.edu.tw//handle/246246/253788錦鯉疱疹病毒（Koi herpesvirus, KHV）目前被分類為水生動物疱疹病毒科（Alloherpesviridae）下的鯉科第三型疱疹病毒（Cyprinid herpesvirus 3, CyHV-3）。自1998年高致死率的錦鯉疱疹病毒傳播開後，嚴重打擊世界各地食用鯉魚以及錦鯉的繁養殖業，台灣也在2002年12月檢疫為錦鯉疱疹病毒疫區。由於目前台灣地區尚未建立適合的細胞可供錦鯉疱疹病毒分離使用，使得台灣的錦鯉疱疹病毒 研究停滯不前。本實驗建立了兩株錦鯉鰭細胞株，分別命名為錦鯉鰭細胞株1（KF1）及錦鯉鰭細胞株2（KF2）並且進行特性分析。兩株錦鯉鰭細胞株皆為表皮樣細胞，並且可生長在含有5-20% 胎牛血清的Leibovitz’s L-15培養基中，最適培養溫度為28-31 ℃，染色體核型均為同源染色體，染色體數目分佈以48居多，並且皆已穩定繼代40代以上。以pEGFP-N1質體DNA轉染錦鯉鰭細胞株，可觀察到綠色螢光蛋白之表現，表示細胞株有潛力利用在表現外來基因的研究。除此之外KF2細胞經過UV照射後在培養的第4小時可觀察到細胞凋亡的現象，未來也可利用於細胞凋亡的研究上。錦鯉鰭細胞株KF2細胞株對錦鯉疱疹病毒具有感受性，其感染力價可達101.5 TCID50/ml，定序後經過序列比對發現與錦鯉疱疹病毒美國株胸腺嘧啶激酶（Thymidine kinase, TK）基因相似度為100%。以上結果顯示所建立的錦鯉鰭細胞株可做為錦鯉疱疹病毒之細胞學研究材料。Koi herpesvirus (KHV) tentatively categorized as a member of the Alloherpesviridae under the species name Cyprinid herpesvirus 3 (CyHV-3). Since 1998, lethal infections of KHV have been spreading, threatening the worldwide production of common carp and koi (both Cyprinus carpio), the first case in Taiwan has been identified in December 2002. Until now, KHV research has impeded by the lack of suitable cell line in Taiwan. In this study, two cell line were established and characterized from koi Cyprinus carpio koi fin tissue. These cell lines have been designated as Koi Fin cell 1 (KF1) and Koi Fin cell 2 (KF2), respectively. Both cell lines were epithelial-like cells. These cells multiplied well in Leibovitz’s L-15 medium, supplemented with 5 to 10% foetal bovine serum, at temperatures between 25 to 31 ℃, and have been subcultured more than 40 passages. Chromosome morphologies of KF1 and KF2 were homogeneous, and the chromosome number of KF1 and KF2 distributed major in 48. These cell lines upon transfection, using pEGFP-N1, produced significant fluorescent signals indicating their potential utility for exogenous studies. Furthermore, UV-irradiated KF2 underwent apoptosis after incubation 4 hr indicating this cell line have potential utility for apoptosis researches. KF2 was susceptible to KHV, and the infection titer can reach 101.5 TCID50/ml, and the sequence alignment is 100% similarity with KHV-USA Thymidine kinase gene. All these results suggest that the established Koi Fin (KF) cell line is suitable for the cell biology study of Koi herpesvirus.2349672 bytesapplication/pdfen-US細胞株建立錦鯉特性分析錦鯉?疹病毒病毒分離Cell lineCyprinus carpio koiCharacterizationCyHV-3Virus isolationthesishttp://ntur.lib.ntu.edu.tw/bitstream/246246/253788/1/ntu-99-R97b45009-1.pdf