Development of a highly efficient system for assessing recombinant gene expression in plant cell suspensions via Agrobacterium tumefaciens transformation

• Published in: Biotechnology and applied biochemistry Vol. 39; no. Pt 3; pp. 355 - 361
• Main Authors: Fuentes, A, Ramos, P.L, Ayra, C, Rodriguez, M, Ramirez, N, Pujol, M
• Format: Journal Article
• Language: English
• Published: United States 01.06.2004
• Subjects: Agrobacterium tumefaciens - genetics; Antibodies, Monoclonal - biosynthesis; Antibodies, Monoclonal - genetics; Antibodies, Monoclonal - immunology; Antigens, Viral - immunology; Bacillus thuringiensis - genetics; Blotting, Western; Caulimovirus - genetics; cell suspension culture; Cells, Cultured; Coculture Techniques; Gene Expression Regulation, Viral; Genes, Bacterial; Glucuronidase - genetics; Glucuronidase - metabolism; Hepatitis B Surface Antigens - biosynthesis; Hepatitis B Surface Antigens - genetics; Hepatitis B Surface Antigens - immunology; Nicotiana - cytology; Nicotiana - genetics; Plants, Genetically Modified - virology; Plants, Toxic - cytology; Plants, Toxic - genetics; Promoter Regions, Genetic; Recombination, Genetic; Solanum lycopersicum - virology; Transformation, Genetic; transgenic plants
• ISSN: 0885-4513; 1470-8744
• DOI: 10.1042/BA20030192

A transient gene-expression system was developed and used to characterize promoter strength, to verify suitability of bacterial gene modifications for expression in plant cells, and to express active antibody molecules. The system is based on suspension tobacco cells transformed by Agrobacterium in a transient way. Conditions such as pre-culture of tobacco cells and the co-cultivation period were identified as determinants to achieve high expression levels. Under established conditions the activity strength of CaMV (cauliflower mosaic virus) 35 S and ToMoTV (tomato mottle taino virus) AL1 promoters were compared. A modified cry gene sequence from Bacillus thuringiensis was expressed and detected by Western-blot analysis. A monoclonal antibody against anti-(hepatitis B virus surface antigen) was produced in such quantities as to allow testing of biological activity and preliminary characterization.