[Construction of human bone morphogenetic protein-2 expressing eukaryotic vector].
OBJECTIVE
To construct human bone morphogenetic protein-2(hBMP-2) expressing eukaryotic vector and observe whether it can be expressed in eukaryotic cells.


METHODS
pUC19 was digested with Sal I and Xba I. The resulting Sal I-Xba I fragment (1.24 kb) which contains the full length of human BMP-2 cDNA was separated on agarose gel and ligated into eukaryotic expression vector pcDNA3 digested with XhoI and XbaI. The recombinant pcDNA3-hBMP-2 plasmid was transferred into fibroblasts cell line NIH3T3. The stable expression of hBMP-2 in the positive cells G418 selected was determined by in situ hybridization and immunohistochemical analysis.


RESULTS
The two fragments digested from recombinant pcDNA3-hBMP-2 plasmid by EcoR I and Xba I represented 1.3 kb and 5.38 kb respectively by agarose electrophoresis, meanwhile the Xho I site was disappeared in pcDNA3-hBMP-2 indicating the successful construction of recombinant pcDNA3-hBMP-2 plasmid. Stable expression of hBMP-2 in pcDNA3-hBMP-2 transfected cells was confirmed by in situ hybridization and immunohistochemical analysis.


CONCLUSIONS
hBMP-2 expressing eukaryotic vector is successfully constructed and can be expressed in eukaryonic cells.
Li
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XD
Institute of Orthopaedics and Traumatology, Xijing Hospital, Fourth Military Medical University, Xi'an Shanxi, P.R. China 710032.
Hu
Y Y
YY
Pu
Q
Q
D016475:3T3 Cells; D000818:Animals; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000096:biosynthesis* / Q000235:genetics; D002478:Cells, Cultured; D005057:Eukaryotic Cells / Q000378:metabolism; D006801:Humans; D051379:Mice; D011994:Recombinant Proteins; D014162:Transfection; D016212:Transforming Growth Factor beta
C525718:BMP2 protein, human; C525719:Bmp2 protein, mouse; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; C499469:recombinant human bone morphogenetic protein-2