Efficient Transfection of Insect Cells with Baculovirus DNA Using Electroporation

• Published in: Journal of general virology 1989-12, Vol.70 (12), p.3501-3505
• Main Authors: Mann, Susan G, King, Linda A
• Format: Article
• Language: English
• Subjects: Animals; Autographa californica; Biological and medical sciences; Biotechnology; cell culture; Cell Line; Cell Survival; DNA, Viral - genetics; electroporation; Fundamental and applied biological sciences. Psychology; Genetic engineering; Genetic technics; Insect Viruses - genetics; Methods. Procedures. Technologies; Microbiology; Moths; plasmids; Techniques used in virology; Transfection; Vectors (cloning, transfer, expression). Insertion sequences and transposons; Virology
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/0022-1317-70-12-3501

School of Biological and Molecular Sciences, Oxford Polytechnic, Gypsy Lane, Headington, Oxford OX3 0BP, U.K. Efficient transfection of Spodoptera frugiperda cells with Autographa californica nuclear polyhedrosis virus (AcNPV) DNA has been carried out using the technique of electroporation. The efficiency of transfection was monitored by assaying the extracellular virus in cell culture supernatants 3 days post-electroporation. Maximum titres of 2 x 10 9 p.f.u./ml AcNPV were obtained when using a pulse length of 7.7 ms and a field strength of 500 V/cm. This compared with a titre of 2 x 10 6 p.f.u./ml AcNPV using the standard calcium phosphate transfection method. Cotransfections of wild-type AcNPV DNA and the transfer plasmid pAcRP23-lacZ were also performed using electroporation and gave -galactosidase recombinant virus titres of 5 x 10 4 p.f.u./ml; this compared with 5 x 10 2 p.f.u./ml using the calcium phosphate method. The maximum proportion of recombinant virus, 2.9%, was obtained by harvesting the transfection medium after 2 days, and using a pulse length of 2.8 ms and a field strength of 750 V/cm. We therefore conclude that electroporation provides a very efficient method for the transfection of insect cells with baculovirus DNA. Keywords: baculovirus DNA, transfection, electroporation Received 15 March 1989; accepted 29 August 1989.