Construction of a lentiviral vector carrying TRAIL gene and its infection efficiency to lymphoma cells in vitro.
- Author:
Xiao-Rui FU
1
;
Xu-Dong ZHANG
;
Chen ZHANG
;
Lu ZHAO
;
Bian-Hong WANG
;
Ming-Zhi ZHANG
Author Information
1. Department of Oncology, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.
- Publication Type:Journal Article
- MeSH:
Base Sequence;
Cell Line, Tumor;
DNA, Complementary;
genetics;
Gene Expression;
Genetic Vectors;
Humans;
Lentivirus;
genetics;
Lymphoma;
genetics;
Molecular Sequence Data;
Plasmids;
TNF-Related Apoptosis-Inducing Ligand;
genetics
- From:
Journal of Experimental Hematology
2012;20(4):900-905
- CountryChina
- Language:Chinese
-
Abstract:
This study was purposed to construct a lentiviral vector carrying the TNF-related apoptosis-inducing ligand (TRAIL) gene and investigate its infection efficiency to several lymphoma cells lines. A pGM-T-TRAIL vector was constructed by inserting the cDNA segment derived from TRAIL mRNA into the cloning vector pGM-T, which was then inserted into the lentiviral vector pWPI. The recombinant lentiviral vector plenti-TRAIL was produced by transfecting 293T cells with pWPI-TRAIL, packaging plasmid Δ8.2, and envelope plasmid pCMV-VSVG and then harvested from the culture supernatant. Infection efficiency was measured in several lymphoma cell lines by live cell GFP fluorescence, while TRAIL expression was assessed by RT-PCR and Western blot. The results showed that the enzyme cut identification and sequencing demonstrated the successful construction of both pGM-T-TRAIL and pWPI-TRAIL. The results of testing drop showed that the concentration of the restructured lentiviral plenti-TRAIL reached 10(9) IU/ml. Comparison of infection efficiency revealed that YTS cells were more likely to be infected than DOHH2 or Jurkat cells (P < 0.05). Finally, RT-PCR and Western blot showed that lymphoma cells infected with plenti-TRAIL were able to efficiently express the TRAIL mRNA and protein. It is concluded that the lentiviral vector pWPI-TRAIL is successfully constructed and the recombinant lentiviral plenti-TRAIL is manufactured. The plenti-TRAIL vector is able to infect several lymphoma cell lines, and the infected lymphoma cells can effectively express TRAIL genes.
