Adaptable doxycycline-regulated gene expression systems for Drosophila.
We have engineered two new versions of the doxycycline (dox) inducible system for use in Drosophila. In the first system, we have used the ubiquitously expressed Drosophila actin5C promoter to express the Tet-Off transactivator (tTA) in all tissue. Induction of a luciferase target transgene begins 6 h after placing the flies on dox-free food. Feeding drug-free food to mothers results in universal target gene expression in their embryos. Larvae raised on regular food also show robust expression of a target reporter gene. In the second version, we have used the Gal4-UAS system to spatially limit expression of the transactivator. Dox withdrawal results in temporally- and spatially-restricted, inducible expression of luciferase in the adult head and embryo. Both the actin5C and Gal4-UAS versions produce more than 100-fold induction of luciferase in the adult, with virtually no leaky expression in the presence of drug. Reporter gene expression is also undetectable in larvae or embryos from mothers fed dox-containing food. Such tight control may be due to the incorporation of Drosophila insulator elements (SCS and SCS') into the transgenic vectors. These systems offer a practical, effective alternative to currently available expression systems in the Drosophila research community.
Stebbins
M J
MJ
Cold Spring Harbor Laboratory, 1 Bungtown Road, Cold Spring Harbor, NY 11724, USA.
Yin
J C
JC
D000199:Actins / Q000235:genetics; D000818:Animals; D030801:Animals, Genetically Modified; D004305:Dose-Response Relationship, Drug; D004318:Doxycycline / Q000494:pharmacology*; D004330:Drosophila / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D004625:Embryo, Nonmammalian / Q000187:drug effects / Q000378:metabolism; D018628:Gene Dosage; D005786:Gene Expression Regulation / Q000187:drug effects; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D007700:Kinetics; D007814:Larva / Q000187:drug effects / Q000378:metabolism; D008156:Luciferases / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics; D019076:Transgenes / Q000235:genetics
D000199:Actins; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D008156:Luciferases; D004318:Doxycycline