Novel vectors for simultaneous high-level dual protein expression in vertebrate and insect cells by recombinant baculoviruses.
Gene transfer into cells of mammalian, avian or piscine origin by baculoviruses carrying expression cassettes active in vertebrate cells (BacMam method) is an attractive alternative to chemical or physical transfection methods or to the use of vectors originating from viruses of vertebrates. For simultaneous high-level expression of two proteins from recombinant baculoviruses we constructed novel dual expression vectors containing human and murine cytomegalovirus immediate-early enhancer/promoter elements in combination with the baculoviral polyhedrin and p10 promoters for simultaneous expression in vertebrate and insect cells. Transduction of ruminant cells with BacMam viruses containing the green fluorescent protein open reading frame downstream from the respective enhancer/promoter elements revealed that a dual expression cassette combining the murine cytomegalovirus immediate-early 1 sequence with the immediate early enhancer/promoter of human cytomegalovirus yields high levels of protein from both transcription units. Protein expression directed by several cytomegalovirus/baculovirus hybrid promoters proceeded efficiently in insect cells infected with the respective recombinants. However, for expression in vertebrate cells the murine ie1 enhancer/promoter upstream the baculoviral p10 promoter was most efficient.
Keil
Günther M
GM
Friedrich-Loeffler-Institut, Greifswald-Insel Riems, Germany. Guenther.Keil@fli.bund.de
Klopfleisch
Constanze
C
Giesow
Katrin
K
Blohm
Ulrike
U
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D002460:Cell Line; D003587:Cytomegalovirus / Q000235:genetics; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007313:Insecta; D018146:Muromegalovirus / Q000235:genetics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D014161:Transduction, Genetic; D014714:Vertebrates
D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins