[Construction of lentivirus vector containing human LIM mineralization protein-1 (LMP-1) and its expression in rat bone mesenchymal stem cells].
OBJECTIVE
To construct a recombiant lentivirus vector of human LMP-1 and detect the expression of LMP-1 in infected rat bone mesenchymal stem cells.


METHODS
LMP-1 gene from the cDNA library were extracted by Polymerase Chain Reaction (PCR). The LMP-1 genes were connected into lentiviral vectors pGC-FU-EGFP which was linearized by Age I enzyme to produce recombiant lentivirus vector called as pGC-FU-LMP-1-EGFP,then packaged by 293T cells. The virus supernant congtaining LV-LMP-1-EGFP was harvested, concentrated and titrated. The rat BMSCs were transfected with recombiant lentivirus LV-LMP-1-EGFP at the most appropriate MOI. The mRNA and protein expression of LMP-1 were detected by RT-PCR and Western blot.


RESULTS
1LV-LMP-1I-EGFP was recombined successfully and the titer reached 2x108TU/ml. 2The efficiency of infection was 93.5% ,which was get after LV-LMP-1-EGFP infected rat BMSCs at the most appropriate MOI=100. The expression of LMP-1 gene was confirmed by RT-PCR and Western blot.


CONCLUSIONS
Lentivirus vector containing human LMP-1 gene is constructed successfully,which can transfected efficiently into rat BMSCs,and the infected rat BMSCs can effectively express LMP-1.
Department of Orthopaedics The Second Hospital of Shanxi Medical University Taiyuan 030001 Shanxi China.
Hui-ming
HM
Hou
Department of Orthopaedics The Second Hospital of Shanxi Medical University Taiyuan 030001 Shanxi China.
Chuan
C
Xiang
Department of Orthopaedics The Second Hospital of Shanxi Medical University Taiyuan 030001 Shanxi China.
Li
L
Guo
Department of Orthopaedics The Second Hospital of Shanxi Medical University Taiyuan 030001 Shanxi China.
Hua-dong
HD
Zhang
D000818:Animals; D005260:Female; D005822:Genetic Vectors; D060850:LIM-Homeodomain Proteins / Q000235:genetics*; D016086:Lentivirus / Q000235:genetics*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014157:Transcription Factors / Q000235:genetics*; D014162:Transfection
C085473:LHX1 protein, human; D060850:LIM-Homeodomain Proteins; D014157:Transcription Factors