摘要
目的构建稳定表达荧光素酶的人乳腺癌MCF-7细胞株。方法采用脂质体转染法将荧光素酶基因导入MCF-7细胞,经潮霉素B筛选建立稳定表达荧光素酶的MCF-7-Luc细胞株;将不同数量MCF-7-Luc细胞分别接种于黑色96孔板(细胞数量59~30000)和裸鼠皮下(细胞数量2.44×10^(3)~5×10^(6)),IVIS LuminaⅢ成像系统体外培养细胞以及裸鼠体内荧光素酶表达效率。结果筛选得到稳定表达荧光素酶的MCF-7-Luc细胞;IVIS LuminaⅢ成像系统检测结果表明,不论是体外培养的细胞或是裸鼠体内,生物发光强度与接种的细胞数量之间均存在明显的线性关系(体外y=129.32x-26409,R^(2)=0.9922,P<0.05;体内y=106.82x+1E+07,R^(2)=0.9404,P<0.05)。结论成功构建了稳定表达荧光素酶的MCF-7-Luc细胞,为研究乳腺癌的发生发展、肿瘤微环境以及抗乳腺癌药物的筛选提供必要条件。
Objective To establish MCF-7 cell strains stably transfected with Luciferase gene(Luc).Methods MCF-7 cells were transfected by lipidosome containing luciferase gene and screened by hygromycin B for establishing MCF-7-Luc lines that could stably express the luciferase.MCF-7-Luc cells were inoculated into the 96-well plate(59~30,000)or inoculated subcutaneously to nude mice(2.44×10^(3)~5×10^(6)),and the expression of reporter gene of luciferase was detected by IVIS LuminaⅢBiological Live Imaging System.Results MCF-7-Luc cells that stably expressed luciferase were screened out.Bioluminescence intensity was correlated in a linear manner with the number of MCF-7-Luc cells in vitro or in vivo(for in vitro correlation,y=129.32x-26409,R^(2)=0.9922,P<0.05;for in vivo correlation,y=106.82x+1 E+07,R^(2)=0.9404,P<0.05).Conclusion MCF-7 cell lines with stable expression of luciferase were established.It will provide conditions for the study of the occurrence and development of breast cancer,and the development anti-breast cancer drugs.
作者
夏敏杰
缪建成
胡晶莹
王玉柱
田芳
杨明俊
杨威
李卫华
王晓东
XIA Min-jie;MIAO Jian-cheng;HU Jing-ying;WANG Yu-zhu;TIAN Fang;YANG Ming-jun;YANG Wei;LI Wei-hua;WANG Xiao-dong(NHC Key Lab of Reproduction Regulation(Shanghai Institute for Biomedical and Pharmaceutical Technologies),Fudan University,Shanghai 200032,China;Laboratory animal Operation Department of Shanghai Institute for Biomedical and Pharmaceutical Technologies 9 Shanghai 201203,China;Shanghai Laboratory Animal Research Center,Shanghai 201203,China)
出处
《毒理学杂志》
CAS
CSCD
2022年第2期142-146,共5页
Journal of Toxicology
基金
国家科技重大专项(2018ZX10301403-005-001)
国家重点研发计划(2016YFC1000905)
上海市第四轮公共卫生三年行动计划重点学科建设计划—环境卫生学与劳动卫生学(15GWZK0201)
上海市科委实验动物研究专项(15140901200)
上海市计划生育科学研究所青年创新基金(Q2016-4)
上海市计划生育科学研究所攀登计划(PD2017-8)。