稳定表达表皮生长因子受体293细胞系的建立及其生物学特性研究

目的建立稳定表达野生型表皮生长因子受体(EGFR)全长的293细胞系。方法通过脂质体瞬间转染的方法将含有EGFR全长的质粒转染293细胞系。利用抗生素筛选的方法使转染阳性的细胞扩增,单克隆化,进一步稳定传代。通过Western印迹的方法挑选高表达克隆,进而使用MTT法测细胞增殖活性。反复传代、冻存、复苏鉴定表达的稳定性。结果获得1株稳定表达EGFR的293细胞系,命名为WT293,且转染前后细胞生长速度存在差异。结论pcDNA3.1(-)载体能够作为稳定表达的载体用于筛选。稳定表达EGFR的293细胞系为配体受体相互作用及靶向EGFR的抗肿瘤药物作用机制的研究提供了实验素材和理论依据。

Establishment of 293 cell line with stable expression of EGFR and its biological character

Objective To establish a 293 cell line that stably expresses wild type epidermal growth factor receptor(EGFR). Methods 293 cell line was transfected by liposome with a plasmid containing whole length EGFR, and positive clones were selected by antibiotic. Western blotting was used to examine the level of EGFR expression, and the proliferation of the cell line was determined by MTT. Results One 293 cell line that stably expressed EGFR was obtained, named WT293, and there was difference of cell proliferation between pre-and post-transfection.Conclusion The vector pcDNA3.1(-)can be used with antibiotic selection to establish stable cell line, which can be used to study the co-effect of ligand and receptor of EGFR, and to elucidate the mechanism of cancer targeted therapy of EGFR.