碩士 / 國立澎湖科技大學 / 水產資源與養殖研究所 / 104 / The establishment and partial characterization of two continuous cell lines from Lethrinus nebulosus are described. Two new continuous cell lines designated SES and SEG were established from snout and gill tissue of spangled emperor, Lethrinus nebulosus respectively. Since their initiation in March 2014, primary cells were cultured in Leibovitz's L-15
medium with 20％ fetal bovine serum (L15-20) at 28℃ and they have been subcultured for more than 100 passages. Before 20 passages, they were subcultured by conditioned medium. After 21 passages, SES were subcultured in L15-7.5 and SEG were subcultured in L15-10. Analysis by light microscopy or scanning electron microscopy was observed that the morphology of SES were fibroblastic and SEG were epithelial. The two continuous cell lines grew well in the temperature range of 18-33℃ and the optimal growth temperature was 28℃. Likewise, the growth rate of the cells increased when the FBS proportion increased from 2.5% to 20%, with the optimal growth at the concentrations of 20% FBS. SES and SEG were gradually cryopreserved by 10% DMSO. They were thawed rapidly and recovered after three months of storage in liquid nitrogen(-196℃). Earliest cells after thawing grew slowly ,but they returned to normal after subculture. Mitochondrial cytochrome b gene fragments of SES,SEG and dorsal muscle of spangled emperor were amplified by polymerase chain reaction(PCR). The PCR products were sequenced and subsequent partial comparative analysis of the identified sequences revealed a 99% match for
SES,SEG and spangled emperor muscle tissue, as well as a 99% match to known spangled emperor mitochondrial DNA sequences (GenBank accession no. AF381269.1). SES cells were susceptible to nervous necrosis virus (NNV), redsea bream iridovirus (RSIV) and hard clam reovirus (HCRV); furthermore, SEG cells were susceptible to RSIV and HCRV.
| Identifer | oai:union.ndltd.org:TW/104NPHT0833001 |
| Date | January 2015 |
| Creators | Chin-Ming Chen, 陳志民 |
| Contributors | Chen-Chun Ku, 古鎮鈞 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 68 |
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