摘要
目的:建立稳定表达萤火虫荧光素酶的人肺癌细胞系并进行体外验证。方法:亚克隆萤火虫荧光素酶基因到真核表达载体pRc/CMV2上,构建重组质粒pRc/CMV2-luc+,转染到肺癌细胞株spc-a-1,经过G418筛选获得单细胞抗性克隆。抗性克隆连续传代,并通过荧光素酶活性检测筛选出高水平表达荧光素酶的稳定细胞克隆,在体外检测细胞的生物发光能力与细胞数量的相关性。结果:构建的pRc/CMV2-luc+真核表达质粒转染spc-a-1细胞后,经G418筛选出多个抗性克隆;传代至40代时确定有3株细胞的荧光素酶活性最高;活体影像系统体外检测证实阳性细胞株发光强度与数量呈正相关。结论:结果表明成功构建了稳定表达萤火虫荧光素酶的spc-a-1细胞系。
Objective：To establish a human lung cancer cell lines with stably expressing firefly luciferase.Methods：Firefly luciferase gene was cloned into vector pRc/CMV2,and recombinant plasmid pRc/CMV2-luc＋ were transfect into spc-a-1 cells by lipofectmine 2000,then spc-a-1 cell clones were obtained by screening with G418.The positive clones stably expressing firefly luciferase were identified from anti-clones by the detection of luciferase activity.Bioluminescence of positive clones were further analyzed in vitro using an in vivo imaging system.Results：When anti-clones were passaged to 40th passage,3 positive clones with high level expression of luciferase were identified.Photons detected with in vivo imaging system were correlated to the numbers of positive clones.Conclusions：spc-a-1 cell lines with stable expression of luciferase gene were successfully established.
出处
《现代生物医学进展》
CAS
2010年第9期1658-1660,共3页
Progress in Modern Biomedicine
基金
广东省自然科学基金(06301124)
广州市属高校科技项目(61093)
