文章摘要
宽口裂腹鱼中肾组织细胞系建立的初步研究*
Preliminary Study on the Establishment of Kidney Tissue Cell Lines in Schizothorax eurystomus
投稿时间:2020-05-12  修订日期:2020-05-31
DOI:
中文关键词: 宽口裂腹鱼  中肾组织  细胞系  细胞培养
英文关键词: Schizothorax eurystomus  Kidney tissue  Cell line  Cell culture
基金项目:国家自然科学基金(31560721, 31860729)、新疆生产建设兵团中青年科技创新领军人才计划项目(2018CB033)和兵团塔里木畜牧科技重点实验室开放课题(HS201804)共同资助[This work was funded by National Natural Science Foundation of China (31560721, 31860729), the Scientific and Technological Innovation Commanding Troops Talented Person Project financially supported by Xinjiang Production and Construction Corps for Young and Middle-aged (2018CB033) and the Key Laboratory of Animal Science and Technology of Xinjiang Production and Construction Corps (HS201804)].
作者单位E-mail
代金彩 1.塔里木大学动物科学学院 新疆 阿拉尔 2.新疆建设兵团塔里木畜牧科技重点实验室 新疆 阿拉尔 1477496028@qq.com 
刘洁雅 新疆建设兵团塔里木畜牧科技重点实验室 新疆 阿拉尔
塔里木大学动物科学学院 新疆 阿拉尔 
15292323133@163.com 
聂竹兰 新疆建设兵团塔里木畜牧科技重点实验室 新疆 阿拉尔
塔里木大学动物科学学院 新疆 阿拉尔 
 
洪继彪 新疆建设兵团塔里木畜牧科技重点实验室 新疆 阿拉尔
塔里木大学动物科学学院 新疆 阿拉尔 
 
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中文摘要:
      鱼类细胞培养是进行种质保存、基因功能分析、细胞工程育种等研究的重要材料和模型。因此对于濒危的宽口裂腹鱼而言,建立细胞系有着极其重要的意义。为了建立宽口裂腹鱼中肾组织细胞系,本文以其中肾组织为材料,以组织块移植法启动细胞原代培养,待细胞稳定后再进行传代培养,建立宽口裂腹鱼中肾组织细胞系,命名为EUM10。分析宽口裂腹鱼中肾细胞系冻存复苏能力和最适生长条件,且通过PCR技术检验污染状况,用线粒体基因片段分析鉴定细胞的来源。结果表明:宽口裂腹鱼中肾组织细胞的最佳培养基为DME/F-12,最适温度25℃,最适血清浓度为20%;细胞呈悬浮生长,生长曲线为“S”型曲线,第10代中肾组织细胞系细胞的群体倍增时间为23.26h;第6代细胞液氮冷冻保存6个月后,复苏存活率达91.91%;进行污染鉴定无污染现象;EUM10第10代细胞线粒体16srRNA序列分析结果与塔里木裂腹鱼基因序列一致性为99.81%,证明细胞系来自宽口裂腹鱼。本文成功建立了宽口裂腹鱼肾组织细胞系,可以将宽口裂腹鱼中肾细胞系作为体外体系运用于宽口裂腹鱼各方面的研究,如细胞遗传学、基因功能分析等,为宽口裂腹鱼的基础研究和育种工作提供基础研究资料。
英文摘要:
      Fish cell culture is an important material and model for germplasm preservation, gene function analysis and cell engineering breeding. Therefore, it is great significance to establish cell lines for endangered Schizothorax eurystomus. In order to establish the kidney tissue cell line in S. eurystomus. In this study, the mesonephric tissue was used as the materialthe, primary cell culture was initiated by tissue block transplantation, Subculture was carried out after the cells were stabilized. A kidney cell line was established, named EUM10. To analyze the cryopreservation and resuscitation ability of renal cell lines and the optimal growth conditions in fish with S. eurystomus, and PCR was used to detect the contamination, mitochondrial gene fragment analysis was used to identify the source of the cell. The results showed that the optimal medium for kidney tissue cells in S.eurystomus was DME/F-12, the optimal temperature was 25 ℃, and the optimal serum concentration was 20%; the cells grew in suspension and the growth curve was “S” type, The curve shows that the population doubling time of the kidney tissue cell line cells in the 10th generation is 23.26h; after 6 months of cryopreservation of liquid nitrogen in the 6th generation, the recovery survival rate is 91.91%. The gene sequence identity of the split-lime fish in Tarim is 99.81%, which proves that the cell line is from the wide-mouth split-bellied fish; there is no pollution in the identification of pollution. Studies on the growth characteristics of fish cell culture mainly focus on culture medium, temperature, serum, freezing and resuscitation. Different medium are used for cells culture. Fish cell culture temperature range is wide. Serum concentration has a great influence on cell growth.Pollution is always a threat to cell culture. In this study, the kidney cell line of S.eurystomus was successfully established. Kidney cell lines can be used as in vitro systems to study various aspects of the fish, such as cytogenetics and gene function analysis. It provides the basic research data for the basic research and breeding of S.eurystomus.
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