电转系统介导的荧光素酶稳定表达PANC.pdf

中国细胞生物学学报 Chinese Journal of Cell Biology 2016, 38(7): 786–793 DOI: 10.11844/cj cb.2016.07.0061 电转系统介导的荧光素酶稳定表达PANC-1细胞的 构建及其生物发光成像检测 屠 珏 黄 宇 凌 云 齐月寒 陈民利* (浙江中医药大学动物实验研究中心/ 比较医学中心, 杭州310053) 摘要 在构建稳定表达荧光素酶的人胰腺癌PANC-1细胞的基础上, 建立适合活体成像研究 的胰腺癌动物模型, 为深入研究胰腺癌发病机制打下基础。该研究运用Lonza NucleofectorTM核转染 系统将携带有荧光素酶基因(firefly luciferase, luc)的质粒(GV258)稳定转染PANC-1细胞, 利用嘌呤霉 素筛选出能稳定表达荧光素酶的细胞株PANC-1-LUC, 并应用活体成像系统对其体内外生物发光进 行检测。进一步构建PANC-1-LUC裸鼠皮下移植瘤, 同样方法接种慢病毒载体构建的PANC-1-luc细 胞作为对照, 观察成瘤率、瘤体积, 检测生物发光强度。体内外生物发光检测结果均显示, 光通量 值与细胞数之间呈显著的直线相关(P0.01)。PANC-1-LUC细胞皮下接种裸鼠, 1周成瘤率即达80% 以上, 瘤体积可达平均100 mm3 。观察6周, 裸鼠皮下肿瘤生长符合Gompertzian 曲线。每周的生物发 光检测结果均显示, PANC-1-LUC移植瘤的光强度与瘤体积有显著的线性关系(P0.05)。以上结果 说明, 该文所构建的PANC-1-LUC细胞株能够稳定、长期地表达荧光素酶, 更适合构建活体成像研 究的裸鼠胰腺癌模型。 关键词 Nucleofector核转染系统; 荧光素酶; PANC-1细胞; 生物发光 Establishment of Pancreatic Cancer Cell Line Stably Expressing Luciferase Mediated by Nucleofector System and Detection with Bioluminescence Imaging System Tu Jue, Huang Yu, Ling Yun, Qi Yuehan, Chen Minli* (Laboratory Animal Research Center/Comparative Medical Research Center, Zhejiang Chinese Medical University, Hangzhou 310053, China) Abstract The paper aimed to establish a pancreatic cancer cell line which could stably produce luciferase and suit for constructing xenotransplanted tumor model in nude mice. PANC-1 cells were transfected with plasmid (GV258) with luc gene mediated by Lonza NucleofectorTM System and monoclonal cell line PANC-1-LUC was screened with puromycin. The luciferase expression of PANC-1-LUC cells was detected by bioluminescence imaging system in vitro and in vivo. The results showed that the bioluminescence density of PANC-1-LUC cells correlated significantly to the cell numbers in vitro and