A recombination-efficient baculovirus vector for simultaneous expression of multiple genes.
The baculovirus system is an extremely powerful tool for expression of heterologous genes in eukaryotic environment. A multiple expression vector, pBacUCmP3, was constructed which harbored two copies of the Autographa californica nuclear polyhedrosis virus very late gene promoter and the Drosophila melanogaster 70-kDa heat-shock protein (hsp70) promoter with downstream unique restriction sites for cloning of three independent foreign genes. Co-transfection of pBacUCmP3 with Bsu36I-linearized viral DNA yields recombinant progeny viruses at very high frequencies. The utility of this multiple expression transfer vector was demonstrated using three heterologous reporter genes encoding the beta-subunit of the human chorionic gonadotropin hormone, firefly luciferase and the bacterial beta-galactosidase (beta Gal) enzyme. The expression of reporter genes, monitored at various times post-infection, confirmed that while beta-Gal synthesis was under the transcriptional control of the hsp70 promoter, the beta hCG and Luc proteins were synthesized as a function of polyhedrin promoter activation profile. This vector will be useful for multiple synthesis of proteins at different time points.
Chatterji
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Eukaryotic Gene Expression Laboratory, National Institute of Immunology, New Delhi, India.
Ahmad
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Venkaiah
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Hasnain
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SE
D000818:Animals; D016367:Baculoviridae / Q000235:genetics* / Q000472:pathogenicity; D018997:Chorionic Gonadotropin, beta Subunit, Human / Q000096:biosynthesis / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods; D001517:Coleoptera / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D017930:Genes, Reporter; D005822:Genetic Vectors / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D007313:Insecta; D008156:Luciferases / Q000235:genetics; D011401:Promoter Regions, Genetic; D015202:Protein Engineering / Q000379:methods; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D013997:Time Factors; D001616:beta-Galactosidase / Q000096:biosynthesis / Q000235:genetics
D018997:Chorionic Gonadotropin, beta Subunit, Human; D011994:Recombinant Proteins; D008156:Luciferases; D001616:beta-Galactosidase