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Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 25(11): 1002-4, 2009 Nov.
Article in Chinese | MEDLINE | ID: mdl-19900367

ABSTRACT

AIM: To obtain high level expression of recombinant human truncated osteoprotegerin (TOPG) with higher bioactivity in CHO-DHFR(-) cells. METHODS: The recombinant vector pcDNA3.1/DHFR-TOPG was constructed and transfected into CHO-DHFR(-) cells by the directions of LipofectAMINE 2000 for stable expression. The stable expression cell strains were screened by selective medium IMDM with 50 mL/L FCS, then serially passed in methotraxate (MTX) for gene amplification. The expression were analyzed by ELISA and RT-PCR. At last, the bioactivity analysis was performed in vitro. RESULTS: The expression level of recombinant truncated human OPG was up to 6 mg/L x 72 h, and it had significant suppression effect on the formation of OLC (P<0.05). CONCLUSION: Recombinant truncated human OPG has high expression and bioactivity. The results make it possible for further studying and clinical implying of OPG.


Subject(s)
Osteoprotegerin/chemistry , Peptide Fragments/biosynthesis , Peptide Fragments/pharmacology , Tetrahydrofolate Dehydrogenase/deficiency , Animals , CHO Cells , Cloning, Molecular , Cricetinae , Cricetulus , DNA, Complementary/genetics , Gene Expression , Genetic Vectors/genetics , Humans , Mice , Osteoclasts/cytology , Osteoclasts/drug effects , Peptide Fragments/genetics , Peptide Fragments/isolation & purification , RNA, Messenger/genetics , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction
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