摘要
宽口裂腹鱼为塔里木河水系的特有物种。为建立其尾鳍细胞系,笔者采用组织块法和胰酶消化法相结合,分别用含胎牛血清的DME/F12培养基体外培养尾鳍组织。初步建立宽口裂腹鱼尾鳍细胞系,细胞传代培养至45代。细胞呈悬浮生长,最适培养液为DME/F12,最适胎牛血清质量分数为20%,最适温度为25℃。第10代细胞的群体倍增时间为24.94 h,细胞呈“S”型生长。第6代细胞液氮冷冻保存180 d后复苏,经台盼蓝染色计数,(88.72±0.99)%的宽口裂腹鱼尾鳍细胞系具有活性,复苏后增殖速度快,可正常传代。细菌、真菌、支原体的鉴定未发现污染。第10代细胞线粒体16S rRNA基因测序结果与GenBank中基因序列做一致性对比,宽口裂腹鱼尾鳍细胞系与NC036933.1的一致率达99.91%,从分子水平证明,宽口裂腹鱼尾鳍细胞系来自宽口裂腹鱼。NaCl质量分数为8‰时,宽口裂腹鱼尾鳍细胞系的相对增殖率最高;NaHCO3质量浓度为7 g/L时,宽口裂腹鱼尾鳍细胞系的相对增殖率最高;宽口裂腹鱼尾鳍细胞系的相对增殖率随盐碱度增加呈先升后降趋势。
Schizostomus eurystomus is a unique species in the Tarim River system. Due to the disturbance of human activities, the change of ecological environment, and the influence of its own factors, the population of S. eurystomus has decreased sharply. The establishment of caudal fin cell lines has not been reported so far. In this study, caudal fin tissues were cultured in vitro of DME/F12 medium containing fetal bovine serum(FBS) by tissue mass method and trypsin digestion method, respectively. The caudal fin cell line was preliminarily established and the caudal fin cells were subcultured for 45 generations. The cell culture method was suspension culture, and the optimal culture medium was DME/F12. The optimal concentration of FBS was 20%. The optimum temperature was 25 ℃. Under the optimal conditions, the population doubling time of the 10th generation cells was determined. The population doubling time of S. eurystomus caudal fin cell line was 24.94 h, and the cells showed an “S” type growth. The 6th generation cells were recovered after being cryopreserved in liquid nitrogen for 180 days. By trypan blue staining,(88.72±0.99)% of S. eurystomus caudal fin cell line cells were active. S. eurystomus caudal fin cell line proliferated quickly after resuscitation and was passed normally. The sequencing results of mitochondrial 16S rRNA of the 10th generation cells were consistent with the GenBank gene sequence, and the consistency rate of S. eurystomus caudal fin cell line and NC036933.1 was 99.91%. At the molecular level, the S. eurystomus caudal fin cell line cell line was confirmed to be from S.eurystomus. NaCl salinity of 8‰ led to increase in the proliferation of S. eurystomus caudal fin cell line. The alkalinity of NaHCO3led to increase in the proliferation of S. eurystomus caudal fin cell line at the maximal rate at NaHCO3alkalinity of 7 g/L. The proliferation of S. eurystomus caudal fin cell line was increased first and then decreased with the increase in salinity.
作者
代金彩
李丽
李学涛
魏杰
聂竹兰
DAI Jincai;LI li;LI Xuetao;WEI Jie;NIE Zhulan(Key Laboratory of Tarim Animal Husbandry Science and Technology,Xinjiang Production and Construction Corps,College of Animal Science,Tarim University,Alar 843300,China;College of Life Science,Tarim University,Alar 843300,China)
出处
《水产科学》
CAS
CSCD
北大核心
2023年第1期30-38,共9页
Fisheries Science
基金
国家自然科学基金资助项目(31560721,31860729)
中国海洋大学塔里木大学科研合作联合基金项目(ZHYLH201902)
华中农业大学塔里木大学科研合作联合基金资助项目(TDHNLH201702,2662017PY118)
新疆维吾尔自治区研究生科研创新项目(TDGRI201918)
国家大学生创新创业训练计划项目(202010757007)。
关键词
宽口裂腹鱼
尾鳍组织
细胞培养
盐碱度
Schizothorax eurystomus
caudal fin tissue
cell culture
salinity and alkalinity
