[Construction of lentiviral vector carrying human VE-cadherin gene and expression of VE-cadherin in leukemic cell line Sup-B15].

Abstract

In order to construct a lentiviral vector carrying human VE-cadherin gene, and to express VE-cadherin in Sup-B15 cells, the VE-cadherin gene was amplified by RT-PCR from the human placenta, and then cloned into pCR-Blunt vector. The VE-cadherin DNA fragment was subcloned into pLB vector to generate a lentiviral vector pLB-VEC. Recombinant lentivirus was generated by co-transfection of three-plasmids into 293FT packing cells using lipofectamine 2000. The Sup-B15 cells were transfected by the lentivirus. The post-transfected Sup-B15 cells were observed by microscopy and flow cytometry. Western blot was used to determine the expression of VE-cadherin. The results showed that the VE-cadherin DNA fragment was amplified from human placenta and was cloned into pCR-Blunt vector, the recombinant lentiviral vector pLB-VEC was successfully constructed. High titer lentivirus was prepared by 3-plasmid packing system, and transfected into Sup-B15 cells in vitro effectively. The obviously morphological changes occurred in transfected cells, the expression of VE-cadherin protein could be detected in Sup-B15 cells via flow cytometry and Western blot. It is concluded that the lentiviral vector pLB-VEC carrying human VE-cadherin gene is successfully constructed; VE-cadherin gene is expressed in Sup-B15 cells via lentiviral vector transfection, which provides an optional tool for further study on the mechanism of VE-cadherin controlling leukemia development.

• Authors: Huan-Xin Zhang, Chong Chen, Ling-Yu Zeng, Zhi-Ling Yan, Zhen-Yu Li, Kai-Lin Xu
• Journal: Zhongguo shi yan xue ye xue za zhi (Zhongguo Shi Yan Xue Ye Xue Za Zhi) Vol. 19 Issue 3 Pg. 574-7 (Jun 2011) ISSN: 1009-2137 [Print] China
• PMID: 21729525 (Publication Type: English Abstract, Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Antigens, CD
• Cadherins
• Recombinant Fusion Proteins
• cadherin 5

Topics

• Antigens, CD (genetics)
• Cadherins (genetics)
• Cell Line, Tumor
• Genetic Vectors
• Humans
• Lentivirus (genetics)
• Plasmids
• Recombinant Fusion Proteins (genetics)
• Transfection