细胞培育肉种子细胞永生化诱导研究  

Research on Immortalization Induction of Cell Cultured Meat Seed Cells

作　　者：杨峰 李莹莹[1] 王守伟[1] 刘文婷 李石磊 Yang Feng;Li Yingying;Wang Shouwei;Liu Wenting;Li Shilei(China Meat Research Center,Beijing Institute of Food Science,Beijing 100068)

机构地区：[1]中国肉类食品综合研究中心,北京食品科学研究院,北京100068

出　　处：《中国食品学报》2022年第12期52-62,共11页Journal of Chinese Institute Of Food Science and Technology

基　　金：国家重点研发计划项目(2021YFC2101404);中国工程院战略研究与咨询项目(2022-XY-22);北京市博士后科研活动经费资助项目(2021ZZ142)。

摘　　要：目的:构建永生化种子细胞是解决细胞培育肉种子细胞来源高效、可持续的有效手段。方法:基于猪成肌细胞,通过端粒酶逆转录酶进行诱导载体的构建,核转染到细胞后分别通过免疫荧光标记、定量PCR、染色体核型分析等方法进行细胞诱导验证。结果:构建永生化细胞诱导载体序列完整度高、无突变;成功将载体转入细胞高效表达并获得诱导阳性细胞株1个;转染后染色体核型未发生不可控的突变;同时诱导后的细胞端粒酶逆转录酶表达量显著高于对照组;更重要的是诱导阳性细胞传代次数突破“Hayflick”增殖界限,实现了200以上的传代次数。结论:构建了猪成肌细胞永生化诱导载体,鉴定了细胞诱导后的稳定性,该永生化种子细胞将在细胞培育肉生产中具有广泛的应用前景。Objective:Construction of immortalized seed cells is an efficient and sustainable way to solve the problem of seed cells of cultured meat.Methods:Based on the current industrialized production of cultured meat,the source of seed cells needs to be solved urgently,and it is also a technical bottleneck in the large-scale production of cell-cultured meat.In this study,based on porcine myoblasts,the inducible vector was constructed by telomerase reverse transcriptase,and the cell induction was verified by immunofluorescence labeling,quantitative PCR,and karyotype analysis after nucleofection into the cells.Results:showed that:the construction of the immortalized cell induction vector had high sequence integrity and no mutation;the vector was successfully transferred into the cells and expressed efficiently,and one induction positive cell line was obtained;no uncontrollable mutation occurred in the chromosomal karyotype after transfection;The expression of telomerase reverse transcriptase in the cells was significantly higher than that in the control group;more importantly,the number of passages of induced positive cells exceeded the'Hayflick'proliferation limit,achieving more than 200 passages.Conclusion:we constructed the immortalization-inducing carrier of porcine myoblasts,described the stability of cells after induction,which will have a broad application prospect in the production process of cultured meat.

关 键 词：细胞培育肉 种子细胞 永生化 诱导载体 端粒酶逆转录酶 

分 类 号：TS219[轻工技术与工程—粮食、油脂及植物蛋白工程]