Transfection of osteoprotegerin gene into Cos-7 cells / 实用口腔医学杂志

ABSTRACT

Objective:To transfect human osteoprotegerin( o pg) gene into Cos-7 cells.Methods:The primers were designed b ased on the human opg cDNA sequence,total mRNA was isolated from 293 cells and RT-PCR was performed.The fragment of opg cDNA was inserted into pSecTag 2/B vector and sequenced by automatic sequence analyzer. The recombined plasmid was transfected into Cos-7 cells by Lipofectamine 2000 and OPG protein expressi on in Cos-7 cells was determined by immunohistochemistry and Western blot. Results:The sequence of opg cDNA from 293 cells obtained by RT- PCR was identical to the sequence provided by GenBank [gi33878056]. The fragm ents of the recombinant plasmid digested with Hind Ⅲ,EcoR I and BamH Ⅰ and exa mined by 10 g/L agarose electrophoresis were consistent with predicted size.OPG over-expressing Cos-7 cells was selected and confirmed by immunohistochemistry and Western blot.Conclusion:Human opg gene may be transfect ed into Cos-7 cells.