Targeted gene expression without a tissue-specific promoter: creating mosaic embryos using laser-induced single-cell heat shock.
We have developed a method to target gene expression in the Drosophila embryo to a specific cell without having a promoter that directs expression in that particular cell. Using a digitally enhanced imaging system to identify single cells within the living embryo, we apply a heat shock to each cell individually by using a laser microbeam. A 1- to 2-min laser treatment is sufficient to induce a heat-shock response but is not lethal to the heat-shocked cells. Induction of heat shock was measured in a variety of cell types, including neurons and somatic muscles, by the expression of beta-galactosidase from an hsp26-lacZ reporter construct or by expression of a UAS target gene after induction of hsGAL4. We discuss the applicability of this technique to ectopic gene expression studies, lineage tracing, gene inactivation studies, and studies of cells in vitro. Laser heat shock is a versatile technique that can be adapted for use in a variety of research organisms and is useful for any studies in which it is desirable to express a given gene in only a distinct cell or clone of cells, either transiently or constitutively, at a time point of choice.
Halfon
M S
MS
Biology Department, Yale University, 640 KBT, P.O. Box 208103, New Haven, CT 06520-8103, USA.
Kose
H
H
Chiba
A
A
Keshishian
H
H
D000818:Animals; D030801:Animals, Genetically Modified; D004268:DNA-Binding Proteins; D004330:Drosophila / Q000196:embryology* / Q000235:genetics; D029721:Drosophila Proteins; D004625:Embryo, Nonmammalian / Q000502:physiology; D005656:Fungal Proteins / Q000096:biosynthesis; D018507:Gene Expression Regulation, Developmental / Q000528:radiation effects; D006360:Heat-Shock Proteins / Q000096:biosynthesis; D006358:Hot Temperature; D007834:Lasers; D009030:Mosaicism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D001616:beta-Galactosidase / Q000096:biosynthesis
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D005656:Fungal Proteins; C052130:GAL4 protein, S cerevisiae; C499528:HSP26 protein, S cerevisiae; D006360:Heat-Shock Proteins; C087442:Hsp26 protein, Drosophila; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; D001616:beta-Galactosidase