Study of the quantitative dinamics of heterologous gene transcription in transfected animals cells for bioprocess optimization

• Grant number: 11/08331-0
• Support Opportunities: Regular Research Grants
• Start date: February 01, 2012
• End date: January 31, 2014
• Field of knowledge: Engineering - Chemical Engineering
• Principal Investigator: Carlos Augusto Pereira
• Grantee: Carlos Augusto Pereira
• Host Institution: Instituto Butantan. Secretaria da Saúde (São Paulo - Estado). São Paulo , SP, Brazil
• Associated researchers: Orlando Garcia Ribeiro Filho ; Renato Mancini Astray ; Soraia Attie Calil Jorge

Abstract

The quantitative RT-PCR (qRT-PCR) is an important tool for gene quantification and few studies have focused its use to study, monitor and control of heterologous gene expression dynamics in eukaryotic systems used for recombinant protein production. Most of the studies, in this area, use the qRT-PCR just as an indirect method for evaluating gene expression. Our aim, on the other hand, is to extend the use of qRT-PCR as a tool for a molecular kinetic detailed study of gene transcription dynamics and bioprocess development in view of better productivity.The aim of this work is to analyze by qRT-PCR the expression of the messenger RNA (mRNA) coding for rabies virus glycoprotein (RVGP) using Drosophila melanogaster S2 cells stably transfected and by BHK-21 cells infected with a recombinant Semliki Forest Virus (SFV) carrying the RVGP mRNA.So, the main purpose of this project is to use the qRT-PCR together with ELISA and flow cytometry for: 1. kinetic studies of mRNA synthesis in several conditions of bioprocess in transfected or infected cells looking for an optimization of recombinant protein synthesis bioprocess in different metabolic conditions, establishing parameters for a bioprocess with higher and better production; 2. Standardization of an original methodology for recombinant no infectious viral particles (rSFV) titration which are today important candidates to viral vaccines. (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

SUAREZ-PATINO, SANDRA FERNANDA; MANCINI, RENATO ASTRAY; PEREIRA, CARLOS AUGUSTO; TORRES SUAZO, CLAUDIO ALBERTO; MENDONCA, RONALDO ZUCATELLI; CALIL JORGE, SORAIA ATTIE. Transient expression of rabies virus glycoprotein (RVGP) in Drosophila melanogaster Schneider 2 (S2) cells. Journal of Biotechnology, v. 192, n. A, p. 255-262, DEC 20 2014. (10/08742-8, 11/08331-0, 02/09482-3)

ASTRAY, R. M.; JORGE, S. A. C.; LEMOS, M. A. N.; YOKOMIZO, A. Y.; BOLDORINI, V. L. L.; PUGLIA, A. L. P.; RIBEIRO, O. G.; PEREIRA, C. A.. Kinetic studies of recombinant rabies virus glycoprotein (RVGP) cDNA transcription and mRNA translation in Drosophila melanogaster S2 cell populations. Cytotechnology, v. 65, n. 5, p. 829-838, OCT 2013. (11/08331-0)

SUAREZ-PATINO, SANDRA FERNANDA; MANCINI, RENATO ASTRAY; PEREIRA, CARLOS AUGUSTO; TORRES SUAZO, CLAUDIO ALBERTO; MENDONCA, RONALDO ZUCATELLI; CALIL JORGE, SORAIA ATTIE. Transient expression of rabies virus glycoprotein (RVGP) in Drosophila melanogaster Schneider 2 (S2) cells. Journal of Biotechnology, v. 192, p. 8-pg., 2014-12-20. (11/08331-0, 10/08742-8, 02/09482-3)