Strong and regulated expression of Escherichia coli beta-galactosidase in insect cells with a baculovirus vector.

Abstract

The N-terminal region of the gene encoding polyhedrin, the major occlusion protein of the insect baculovirus Autographa californica nuclear polyhedrosis virus (AcNPV), has been fused to DNA encoding Escherichia coli beta-galactosidase. The fused gene was inserted into the AcNPV DNA genome by cotransfection of insect cells with recombinant plasmid DNA and wild-type AcNPV genomic DNA. Recombinant viruses were selected as blue plaques in the presence of a beta-galactosidase indicator, 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside. Studies of one such virus, L1GP-gal3, indicated that the synthesis of beta-galactosidase is temporally controlled beginning late (20 h) in infection after the release of infectious virus particles from the cell. By 48 h postinfection, a remarkably high level of expression is achieved. On the basis of these results, AcNPV should be a useful vector for the stable propagation and expression of passenger genes in a lepidopteran cell background. A generalized transplacement vector that facilitates the construction and selection of recombinant viruses carrying passenger genes under their own promoter control has also been developed.

• Authors: G D Pennock, C Shoemaker, L K Miller
• Journal: Molecular and cellular biology (Mol Cell Biol) Vol. 4 Issue 3 Pg. 399-406 (Mar 1984) ISSN: 0270-7306 [Print] United States
• PMID: 6325875 (Publication Type: Journal Article)

Chemical References

• DNA Restriction Enzymes
• Galactosidases
• beta-Galactosidase

Topics

• Amino Acid Sequence
• Animals
• Base Sequence
• Cell Line
• DNA Restriction Enzymes
• Escherichia coli (enzymology, genetics)
• Galactosidases (genetics)
• Genes
• Genes, Bacterial
• Genetic Vectors
• Insect Viruses (genetics)
• Lepidoptera
• Plasmids
• beta-Galactosidase (genetics, metabolism)