Document Detail


A rapid and efficient method to express target genes in mammalian cells by baculovirus.
MedLine Citation:
PMID:  15162535     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
AIM: To investigate the modification of baculovirus vector and the feasibility of delivering exogenous genes into mammalian cells with the culture supernatant of Spodoptera frugiperta (Sf9) cells infected by recombinant baculoviruses. METHODS: Two recombinant baculoviruses (BacV-CMV-EGFPA, BacV-CMV-EGFPB) containing CMV-EGFP expression cassette were constructed. HepG2 cells were directly incubated with the culture supernatant of Sf9 cells infected by recombinant baculoviruses, and reporter gene transfer and expression efficiencies were analyzed by flow cytometry (FCM). The optimal transduction conditions were investigated by FCM assay in HepG2 cells. Gene-transfer and expression efficiencies in HepG2 or CV1 cells by baculovirus vectors were compared with lipofectAMINE, recombinant retrovirus and vaccinia virus expression systems. Twenty different mammalian cell lines were used to investigate the feasibility of delivering exogenous genes into different mammalian cells with the culture supernatant of infected Sf9 cells. RESULTS: CMV promoter could directly express reporter genes in Sf9 cells with a relatively low efficiency. Target cells incubated with the 1:1 diluted culture supernatant (moi=50) for 12 h at 37 degrees C could achieve the highest transduction and expression efficiencies with least impairment to cell viability. Under similar conditions the baculovirus vector could achieve the highest gene-transfer and expression efficiency than lipofectAMINE, recombinant retrovirus and vaccinia virus expression systems. Most mammalian cell lines could be transduced with recombinant baculovirus. In primate adherent culture cells the recombinant baculovirus could arrive the highest infection and expression efficiencies, but it was not very satisfactory in the cell lines from mice and suspended culture cells. CONCLUSION: Mammalian cells incubated with the culture supernatant of infected Sf9 cells could serve as a very convenient way for rapid and efficient expression of foreign genes in mammalian cells, but it might be more suitable for primate adherent culture cells.
Authors:
Tong Cheng; Chen-Yu Xu; Ying-Bin Wang; Min Chen; Ting Wu; Jun Zhang; Ning-Shao Xia
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  World journal of gastroenterology : WJG     Volume:  10     ISSN:  1007-9327     ISO Abbreviation:  World J. Gastroenterol.     Publication Date:  2004 Jun 
Date Detail:
Created Date:  2004-05-26     Completed Date:  2004-07-19     Revised Date:  2008-11-21    
Medline Journal Info:
Nlm Unique ID:  100883448     Medline TA:  World J Gastroenterol     Country:  China    
Other Details:
Languages:  eng     Pagination:  1612-8     Citation Subset:  IM    
Affiliation:
Key Laboratory of Cell Biology and Tumor Cell Engineering of Ministry of Education, Xiamen University, Xiamen 361005, Fujian Province, China.
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MeSH Terms
Descriptor/Qualifier:
Animals
Baculoviridae / genetics*
CHO Cells
Carcinoma, Hepatocellular
Cricetinae
Gene Expression
Gene Transfer Techniques*
Genes, Reporter / genetics
Green Fluorescent Proteins
Hela Cells
Humans
Liver Neoplasms
Luminescent Proteins / genetics
Mammals
Mice
NIH 3T3 Cells
Plasmids / genetics*
Promoter Regions, Genetic / genetics
Recombinant Proteins / genetics
Spodoptera
Chemical
Reg. No./Substance:
0/Luminescent Proteins; 0/Recombinant Proteins; 147336-22-9/Green Fluorescent Proteins

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