关键词: 受体, 雌激素,激素非依赖性前列腺癌细胞,前列腺肿瘤,真核表达,遗传载体,转染

Abstract: Objective To construct an eukaryotic expression vector of human ERβ (hERβ) full length gene which named pEGFP-C1-hERβ and transfect it into hormone-independent prostate cancer cell line PC-3M. Methods The complete gene of hERβ（1 593 bp） was obtained with the technique of RT-PCR by using human ovary tissue mRNA as template which obstained from the ovarian cyst patients. The PCR product was connected with pMD18-T vector and sequenced automatically，and then the connected product and the expression vector pEGFP-C1 were digested by restrictive enzyme Hind Ⅲ and BamH Ⅰ， respectively，and the pEGFP-C1-hERβ vector was constructed by using gene recombinant technique. The plasmid was detected by endonuclease digestion and PCR，and then was transfected to PC-3M cells by lipid reagent. Results The clone of the complete hERβ gene and the construction of expression vector were finished successfully by endonuclease digestion and sequenced automatically. The product of the endomuclease digestion was as long as the human complete hERβ gene (1 612 bp)，and sequence analysis suggested that the hERβ sequence detected by PCR was identical to that published in GenBank (NM_001437). The expression of green fluorescence protein (GFP) was observed under the fluorescence microscope. Conclusion The eukaryotic expression vector pEGFP-C1-hERβ is constructed and it expresses in PC-3M cells successfully.

Key words: receptors, estrogen, independent prostate cancer cells, prostatic neoplasms,gene expression, genetic vectors, transfections