人乳腺珠蛋白和亲脂素B共转染乳腺癌细胞T47-D

目的构建人乳腺珠蛋白（Mammaglobin A）和亲脂素B（Lipophilin B）真核表达载体，建立稳定高表达Mammaglobin A和同时稳定高表达Mammaglobin A和Lipophilin B的人乳腺癌T47-D细胞系。方法通过RT-PCR的方法从人乳腺癌细胞系MDA—MB-415中扩增MammaglobinA和LipophilinB基因，构建真核表达载体pc—MA、pc—LP，分别独立及共转染T47-D细胞，G418筛选阳性克隆，并利用RT—PCR、Western Blot鉴定。结果重组质粒pc—MA、pc—LP酶切图谱、序列分析证明pc—MA、pc—LP载体构建成功，目的基因序列与GenBank中的序列完全一致。RT-PCR与Western Blot结果证实，独立稳定高表达Mammaglobin A和同时稳定高表达Mammaglobin A和Lipophilin B的人乳腺癌细胞系建立成功。结论成功建立独立高表达Mammaglobin A和同时高表达Mammaglobin A和Lipophilin B的乳腺癌细胞系，为探讨Maremaglobin A和Lipophilin B在乳腺癌发生发展中的作用提供了实验基础。

Co-transfection of Mammaglobin A and Lipophilin B in breast cancer T47-D cells

Objective To construct the eukaryotic expression recombinant plasmid expressing Mammaglobin A and Lipophilin B and to establish human breast cancer T47-D cell lines stable expression Mammaglobin A and stable co-expression Mammaglobin A and Lipophilin B. Methods The Mammaglobin A and Lipophilin B genes were cloned from human breast cancer cell line MDA-MB-415, the eukaryotic expression vectors pc-MA, pc-LP were constructed and then transfected in to T47-D cells. The positive cells were screening by G418 and examined by RT-PCR and Western Blotting analysis. Results Recombinant plasmids expressed Mammaglobin A and Lipophilin B were successfully constructed confirmed by restriction enzyme digestion and DNA sequencing analysis. Stable expression of ectopic Mammaglobin A and Lipophilin B in T47-D cells were examined by RT-PCR and Western Blot analysis. Conclusion T47-D Cell line which stably expressed ectopic Mammaglobin A and Lipophilin B is successfully established. It provides a model for studying the function of Mammaglobin A and Lipophilin B in the breast carcinogenesis.