Low-light image analysis of transgenic organisms using bacterial luciferase as a marker.
Methods for measurement of a novel light-emitting reporter gene system in bacteria, yeast, plant cells, plant tissues and intact plant organs are described. The principle underlying the assay procedures is the bacterial luciferase catalysed oxidation of reduced flavin mononucleotide (FMNH2) in the presence of the ten carbon aldehyde decanal, to yield FMN, decanoic acid, water and a photon of light at 490 nm which can be captured by X-ray film, a photomultiplier tube or, for in vivo measurements, an image-intensifier coupled to a video camera. This light measuring assay system is sensitive, easy to use, inexpensive, does not require radioactivity, and has been used successfully for rapid detection of bacterial transformants, the quantitative measurement of transient and stable gene expression in bacteria and yeast, and in vivo measurement of temporal and spatial gene expression throughout plant and animal development.
Langridge
W
W
Department of Plant Science, University of Alberta, Edmonton, Canada.
Escher
A
A
Wang
G
G
Ayre
B
B
Fodor
I
I
Szalay
A
A
D000818:Animals; D030801:Animals, Genetically Modified; D018835:Chaperonin 10 / Q000096:biosynthesis; D018834:Chaperonin 60 / Q000096:biosynthesis; D001517:Coleoptera / Q000201:enzymology; D015870:Gene Expression; D005798:Genes, Bacterial; D007313:Insecta; D008156:Luciferases / Q000032:analysis* / Q000096:biosynthesis; D008163:Luminescent Measurements; D030821:Plants, Genetically Modified; D017510:Protein Folding; D014162:Transfection; D014733:Vibrio / Q000201:enzymology
D018835:Chaperonin 10; D018834:Chaperonin 60; D008156:Luciferases