特异性表达新霉素耐药基因Neo^R的人乳腺癌细胞模型的建立

ESTABLISHMENT OF HUMAN BREAST CANCER CELL MODEL EXPRESSING SPECIFIC NEOMYCIN RESISTANCE GENE Neo R

将含有ＮｅｏＲ基因的ｐＳＶ２－ｎｅｏ质粒经扩增、提取及纯化，以磷酸钙共沉淀法转染入ＭＤＡ人乳腺癌细胞系，经Ｇ４１８长期高压筛选出７株抗性ＭＤＡ细胞克隆，采用ＮｅｏＲ基因特异性引物进行聚合酶链反应，证实了ＮｅｏＲ基因在各株Ｇ４１８抗性ＭＤＡ细胞基因组中的整合。ＭＤＡ－ＮｅｏＲ细胞在传代６个月之后仍可耐受６００ｍｇ／Ｌ高浓度的Ｇ４１８，说明ＮｅｏＲ基因导入后的整合及表达是稳定的。表明已初步建立了携有特异标志基因ＮｅｏＲ的人乳腺癌细胞模型。

The pSV2 neo plasmid were cleavaged by EcoR I enzyme after amplification, extraction and purification from E.coli strain, then transfected into MDA cells, a breast cancer cell line, by method of Calcium phosphate DNA precipitate. Seven strains of cell lines of G418 resistance were obtained. The integration of Neo R gene in these G418 resistance MDA cell genome was identified by PCR methods using Neo R specific primers. MDA Neo R cells cultured in vitro for six months are still endurable for G418(600mg/L). So, A breast cancer cell model containing specific Neo R gene has been established.