Dual luminescence-based reporter gene assay for luciferase and beta-galactosidase.
A unique combined luminescence assay for firefly (Photinus pyralis) luciferase and beta-galactosidase (beta-gal) reporter gene products is described. Luciferase and beta-gal activities are determined with the same aliquot of cell lysate prepared from cells contransfected with both reporter genes, thereby reducing manual labor and increasing experimental accuracy. With the Dual-Light assay system, luciferase activity is measured first with an enhanced luciferase assay, followed by quantitation of beta-gal with Galacton-Plus chemiluminescent substrate and Sapphire-II enhancer. Highly sensitive detection of luciferase (2 fg) and beta-gal (8 fg) is achieved with a dynamic range over seven orders of magnitude of enzyme concentration. Comparative analysis of both independent and combined (Dual-Light) detection methods for cells contransfected with luciferase and beta-gal reporter genes is also described.
Martin
C S
CS
Tropix, Inc., Bedford, MA, USA.
Wight
P A
PA
Dobretsova
A
A
Bronstein
I
I
D000818:Animals; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D005822:Genetic Vectors; D008156:Luciferases / Q000032:analysis*; D008163:Luminescent Measurements; D051379:Mice; D008832:Microchemistry; D018991:Myelin Proteolipid Protein / Q000235:genetics; D010783:Photometry / Q000295:instrumentation; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000032:analysis*; D014162:Transfection; D001616:beta-Galactosidase / Q000032:analysis*
D018991:Myelin Proteolipid Protein; D011993:Recombinant Fusion Proteins; D008156:Luciferases; D001616:beta-Galactosidase