Rabies virus glycoprotein expression in Drosophila S2 cells: Influence of re-selection on protein expression

被引：3
作者：
Dos Santos, Alexandra Souza [1 ]
Lemos, Marcos Alexandre Nobre [1 ]
Pereira, Carlos Augusto [1 ]
Jorge, Soraia Attie Calil [1 ]
机构：
[1] Laboratório de Imunologia Viral, Instituto Butantan, São Paulo
来源：
Biotechnology Journal | 2009年 / 4卷 / 11期
关键词：
Hygromycin re-selection; RVGP; S2; cells;
D O I：
10.1002/biot.200900123
中图分类号：
学科分类号：
摘要：
The aim of this study was to achieve expression of recombinant rabies virus glycoprotein (rRVGP) in Drosophila S2 cells. For this, a cDNA coding for the selection hygromycin antibiotic and the cDNA encoding the RVGP protein under the control of the constitutive actin promoter (Ac) were cloned in an expression plasmid, which was transfected into S2 cells. S2 cell populations (S2AcRVGPHy) showed rRVGP expression in cell lysates, attaining concentrations up to 1.5 μg/107 cells (705 μg/L). Of the transfected cells, 20% were shown to express the rRVGP. Cell subpopulations selected by limiting dilution expressed higher rRVGP yields and 90% of the cells were shown to express the rRVGP. Cell populations re-selected by addition of hygromycin were shown to express 10 times higher rRVGP yields. The data presented here show that Drosophila S2 cells can be efficiently transfected with an expression/selection plasmid for rRVGP expression, allowing its synthesis with a high degree of physical and biological integrity. The importance of subpopulation selection was indicated by the increasing rRVGP yields during these procedures. © 2009 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.
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页码：1578 / 1581
页数：3
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共 16 条
  • [11] Perrin P., Lafon M., Sureau P., Enzyme linked immuno-sorbent assay (ELISA) for the determination of glycoprotein content of rabies vaccines, Laboratory techniques in rabies, pp. 383-388, (1996)
  • [12] Santos M.G., Jorge S.A., Brillet K., Pereira C.A., Improving heterologous protein expression in transfected Drosophila S2 cells as assessed by EGFP expression, Cytotechnology, 54, pp. 15-24, (2007)
  • [13] Jorge S.A.C., Santos A.S., Spina A., Pereira C.A., Expression of the hepatitis B virus surface antigen in Drosophila S2 cells, Cytotechnology, 57, pp. 51-59, (2008)
  • [14] Iwaki T., Castellino F.J., A single plasmid transfection that offers a significant advantage associated with puromycin selection in Drosophila Schneider S2 cells expressing heterologous proteins, Cytotechnology, 57, pp. 45-49, (2008)
  • [15] Calos M.P., Lebkowski J.S., Botchan M.R., High mutation frequency in DNA transfected into mammalian cells, Proc. Natl. Acad. Sci. USA, 80, pp. 3015-3019, (1983)
  • [16] Razzaque A., Mizusawa H., Seidman M.M., Rearrangement and mutagenesis of a shuttle vector plasmid after passage in mammalian cells, Proc. Natl. Acad. Sci. USA, 80, pp. 3010-3014, (1983)
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