基于“睡美人”转座系统构建可视化黑色素细胞瘤及肿瘤模型  

Establishment of visual melanoma cell line and animal model based on "Sleeping Beauty" transposon

作　　者：陈慧玲[1] 彭彩虹[2] 李小丽[4] 韩悌云 张德奎[3] Chen Hui-ling;Peng Cai-hong;Li Xiao-li;Han Ti-yun;Zhang De-kui(Department of Hematology,The Second Hospital of Lanzhou University,Lanzhou 730030,China;Department of Medical Oncology,The Second Hospital of Lanzhou University,Lanzhou 730030,China;Key Laboratory of Digestive Diseases,The Second Hospital of Lanzhou University,Lanzhou 730030,China;The Second School of Clinical Medicine,Lanzhou University,Lanzhou 730000,China)

机构地区：[1]兰州大学第二医院血液科,甘肃兰州730030 [2]兰州大学第二医院肿瘤内科,甘肃兰州730030 [3]兰州大学第二医院消化疾病重点实验室,甘肃兰州730030 [4]兰州大学第二临床医学院,甘肃兰州730000

出　　处：《兰州大学学报（医学版）》2022年第1期9-12,共4页Journal of Lanzhou University（Medical Sciences）

基　　金：兰州大学第二医院“萃英科技创新”计划资助项目(CY2019-QN16)。

摘　　要：目的利用"睡美人"(SB)转座系统构建可表达红色荧光蛋白和荧光素酶基因的黑色素瘤B16F10细胞系,建立可视化黑色素瘤动物模型。方法从含荧光素酶Luc2的质粒中获取Luc2基因,用NcoⅠ和XbaⅠ限制性内切酶双酶切后插入pSBbi-RP SB转座子系统。构建的重组pSBbi-Luc2-RP转座质粒与转座酶按照一定比例转染黑色素瘤细胞B16F10,利用荧光显微镜观察转染效率,经嘌呤霉素筛选稳定转染的细胞系。利用活体成像系统可视化实时监测评价小鼠皮下成瘤模型。结果利用SB转座系统成功构建黑色素瘤B16F10细胞系,24、48 h转染率分别达40%、70%,上述细胞系经皮下成瘤后1周活体动物成像系统可检测到肿瘤形成。结论利用SB转座系统可快速建立可视化黑色素瘤细胞系,利用小动物活体成像系统可动态观察黑色素瘤动物模型中肿瘤生长情况。Objective To construct a melanoma cell line B16F10 that can express red fluorescent protein and luciferase gene,based on"Sleeping Beauty"(SB)transposon,and establish a visual melanoma animal model.Methods Luc2 gene was obtained from a plasmid containing luciferase,and inserted into pSBbi-RP SB transposon after digestion with NcoⅠand XbaⅠrestriction endonuclease.The recombinant vector pSBbiLuc2-RP and transposase pSB100X were mixed by a certain proportion and transfected into B16F10 cells.The transfection efficiency was examined by fluorescence microscopy,the stable transfection cell line was screened by purinomycin.In vivo imaging system was used to visually monitor and evaluate the melanoma model.Results The melanoma cell line B16F10 was successfully constructed by SB transposon system.The transfection rates were 40%and 70%at 24 h and 48 h,respectively.Tumor formation of the cell lines could be detected by animal imaging system one week after subcutaneous tumor formation.Conclusion Visualized melanoma cell lines can be rapidly established by SB transposon system,and tumor growth in melanoma animal models can be dynamically observed by using in vivo imaging system.

关 键 词：“睡美人”转座系统 可视化 黑色素瘤 活体成像 

分 类 号：R73-7[医药卫生—肿瘤]