锦鲤鳍条组织细胞系的建立及其生物学特性

采用组织块移植培养技术,对来源于锦鲤（Cryprinus carpiod）鳍条组织的细胞进行原代培养,建立了锦鲤鳍条组织细胞系,已稳定传代60多次,命名为Koi-Fin。锦鲤鳍条组织细胞为成纤维样细胞,最佳培养基为MEME,最适血清体积分数为10%,最适培养温度为25 oC,群体倍增时间为43.5 h。该细胞经液氮冷冻保藏12个月后采用台盼蓝染色,约（80.21±5.84）%的细胞具有细胞活性,复苏细胞生长旺盛。细胞染色体分析显示,第16代锦鲤鳍条组织细胞的染色体数目为正常二倍体2n=100,第40代细胞的染色体众数为52。病毒敏感性试验结果表明,Koi-Fin细胞系对锦鲤疱疹病毒（Koi Herpesvirus,KHV）敏感,可产生典型细胞病变效应,病毒滴度为107.86±0.51TCID50/mL。针对锦鲤疱疹病毒胸苷激酶（thymidine kinase,TK）基因设计特异性引物进行PCR检测,可扩增出病毒靶基因片段。

Establishment and Characterization of a Cell Line Derived from Fin of Cryprinus carpiod

A cell line, designated Koi-Fin, has been established from the fin of Cryprinus carpiod by using tissue explant techniques. Koi-Fin cell line has been continuously subcultured over 60 times and fully characterized in the aspects of morphological observation, cryopreservation, optimal growth kinetics, population doubling time and karyotyping, etc. Koi-Fin cell monolayer consists of fibroblast-like cells and its optimal growth condition is： MEM medium, 10% FBS and 25 ℃ Under such optimized condition the doubling time of Koi-Fin cell num- bers is about 46.5 h. After 12 months cryopreservation in liquid nitrogen, Koi-Fin cells exhibit a viability of about （80.21±5.84）% with trypan blue staining. Chromosome typing of this cell line revealed that at its 16th passage the diploid chromosome number was 2n=100, and at the 40th passage the modal number is 52. The results of virus sen- sitivity test indicated that Koi-Fin cell line is susceptible to the infection of Koi Herpesvirus （KHV）, a characteristic viral-induced cytopathic effect （CPE） was observed and the viral titer （TCIDs0/mL） reached about 10^7.86＋0.51. Moreover, PCR detection demonstrated that the targeted gene can be amplified by using specific primers designed targeting the viral thymidine kinase gene.