| Full text | |
| Author(s): |
Nobre Lemos, Marcos Alexandre
[1]
;
dos Santos, Alexandra Souza
[1]
;
Astray, Renato Mancini
[1]
;
Pereira, Carlos Augusto
[1]
;
Calil Jorge, Soraia Attie
[1]
Total Authors: 5
|
| Affiliation: | [1] Inst Butantan, Lab Imunol Viral, BR-05503900 Sao Paulo - Brazil
Total Affiliations: 1
|
| Document type: | Journal article |
| Source: | Journal of Biotechnology; v. 143, n. 2, p. 103-110, AUG 20 2009. |
| Web of Science Citations: | 16 |
| Abstract | |
The cDNA encoding the rabies virus glycoprotein (RVGP) gene was cloned in expression plasmids under the control of the inductive metallothionein promoter. They were designed in order to bear or not a secretion signal (i) and a cDNA coding for the selection hygromycin. These vectors were transfected into S2 cells, cell populations selected and subpopulations were then obtained by reselection with hygromycin. Cell cultures were examined for kinetics of cell growth, detection of RVGP mRNA and expression of RVGP. All cell populations were shown to express the RVGPmRNA upon induction. S2MtRVGPHy cell population. transfected with one vector that contains RGPV gene and selection gene, was shown to express higher amounts of RVGP as evaluated by flow cytometry (similar to 52%) and ELISA (0.64 mu g/10(7) cells at day 7). Subpopulation selection allowed a higher RVGP expression, specially for the S2MtRVGPHy(+) (5.5 mu g/107 cells at day 7). NaBu treatment leading to lower cell growth and higher RVGP expression allowed an even higher RVGP synthesis by S2MtRVGPHy(+) (8.4 mu g/10(7) cells at day 7). SF900II medium leading to a higher S2MtRVGPHy(+) cell growth allowed a higher final RVGP synthesis in this cell culture. RVGP synthesis maybe optimized by the expression/selection vectors design, cell subpopulations selection, chromatin exposure and culture medium employed. (C) 2009 Elsevier B.V. All rights reserved. (AU) | |
| FAPESP's process: | 02/09482-3 - Heterologus gene expression in dipteran cells: molecular biology and bioprocess engineering |
| Grantee: | Carlos Augusto Pereira |
| Support Opportunities: | Research Projects - Thematic Grants |