Firefly luciferase gene transmission and expression in transgenic medaka (Oryzias latipes).
Plasmids containing the luciferase gene from the firefly (Photinus pyralis) fused to the Chinese hamster metallothioneine I promoter (ChMTI) were microinjected into the pronuclei of medaka (Oryzias latipes) eggs, which were then artificially inseminated. Evidence of integration into the genome was gained from observation of germ-line transmission in a mendelian fashion from the F1 to the F2 generation. However, gene expression (light emission) could not be demonstrated in the established transgenic line. In a separate program, transient expression of gene constructs containing the luciferase gene fused to various promoters was compared in medaka embryos. Plasmids were microinjected into pronuclei, and homogenates from 3-day-old embryos were measured for light emission using a luminometer. Among the various promoters tested (SV40, RSV-LTR, ChMTI, HSP70, and mouse albumin), the highest levels of luciferase gene expression were observed in gene constructs containing ChMTI and HSP70 gene promoters. Expression in these two constructs was significantly increased following administration of ZnSO4 or heat treatment, respectively. Plasmids were also introduced into goldfish fibroblast-like cells in vitro, in which enzymatically active luciferase was transiently expressed. Assaying for expression of luciferase provided a rapid and sensitive method for monitoring promoter activity. The potential usefulness of this fish species for cancer research is discussed based on accumulated information from carcinogenesis studies.
Sato
A
A
Department of Experimental Pathology and Cell Biology, Cancer Institute, Tokyo, Japan.
Komura
J
J
Masahito
P
P
Matsukuma
S
S
Aoki
K
K
Ishikawa
T
T
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics*; D015139:Blotting, Southern; D002478:Cells, Cultured; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D001517:Coleoptera / Q000235:genetics; D004625:Embryo, Nonmammalian / Q000201:enzymology; D005260:Female; D015971:Gene Expression Regulation, Enzymologic; D008156:Luciferases / Q000096:biosynthesis / Q000235:genetics*; D008297:Male; D008845:Microinjections; D009990:Oryzias / Q000235:genetics*; D010375:Pedigree; D011401:Promoter Regions, Genetic / Q000502:physiology; D014162:Transfection
D008156:Luciferases