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KMID : 0390320120220020079
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Chungbuk Medical Journal
2012 Volume.22 No. 2 p.79 ~ p.84
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Preparation of a Lentivirus expressing Mammalian Ste20-like kinase 2
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Lee Yong-Hee
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Abstract
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Purpose: Mammalian ste20-like kinase (MST) signaling pathway is an important dual regulator of apoptosis and cell proliferation and is emerging as a novel tumor suppressor pathway. The aim of this study was to prepare an MST2-lentivirus that can overexpress MST2 protein in cells that are resistant to transfection of foreign genes.
Materials and Methods: MST2 cDNA with 3 HA tags was subcloned into the lentiviral transfer vector, pFUGW which includes a GFP expression cassette. HEK293 cells were transfected with this plasmid to confirm MST2 expression. Lentivirus was prepared by transfecting 293T cells with pFUGW-3HA-MST2, envelope glycoprotein expression vector, and packaging vector.
Results: MST2 expression was confirmed by GFP fluorescence and immunoblotting with HA antibody in HEK293 cells transfected with pFUGW-3HA-MST2. HeLa and A549 cells were transduced with MST2-lentivirus and showed strong GFP fluorescence. Fully differentiated 3T3-L1 adipocytes are difficult to transfect with foreign genes. However, they were successfully transduced with MST2-lentivirus and showed strong MST2 expression.
Conclusion: In this study, we generated and characterized an MST2-expressing lentivirus and this will be a valuable experimental tool for the study of MST kinase signaling pathway.
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KEYWORD
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MST2, Protein kinase, Lentivirus
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