Temporal and spatial control of transgene expression using laser induction of the hsp70 promoter.
BACKGROUND
Precise temporal and spatial regulation of transgene expression is a critical tool to investigate gene function in developing organisms. The most commonly used technique to achieve tight control of transgene expression, however, requires the use of specific DNA enhancers that are difficult to characterize in non-model organisms. Here, we sought to eliminate the need for this type of sequence-based gene regulation and to open the field of functional genetics to a broader range of organisms.


RESULTS
We have developed a new laser mediated method to heat shock groups of cells that provides precise spatio-temporal control of gene expression without requiring knowledge of specific enhancer sequences. We tested our laser-system in a transgenic line of Bicyclus anynana butterflies containing the EGFP reporter gene attached to the heat sensitive hsp70 promoter of Drosophila melanogaster. Whole organismal heat shocks demonstrated that this Drosophila promoter can drive gene expression in butterflies, and the subsequent laser heat shocks showed that it was possible to activate cell-specific gene expression in very precise patterns on developing pupal wings.


CONCLUSIONS
This laser-mediated gene expression system will enable functional genetic investigations, i.e., the ectopic expression of genes and their knock-down in targeted groups of cells in model and non-model organisms with little or no available regulatory data, as long as a compatible heat-shock promoter is used and the target tissue is accessible to a laser beam. This technique will also be useful in evolutionary developmental biology as it will enable the study of the evolution of gene function across a variety of organisms.
Ramos
Diane M
DM
Department of Biological Sciences, University at Buffalo, Buffalo, NY 14260, USA. dmsmith2@buffalo.edu <dmsmith2@buffalo.edu>
Kamal
Firdous
F
Wimmer
Ernst A
EA
Cartwright
Alexander N
AN
Monteiro
Antónia
A
D000818:Animals; D030801:Animals, Genetically Modified / Q000235:genetics; D015139:Blotting, Southern; D002080:Butterflies / Q000235:genetics* / Q000254:growth & development; D004331:Drosophila melanogaster / Q000235:genetics* / Q000254:growth & development*; D018507:Gene Expression Regulation, Developmental; D017930:Genes, Reporter; D018773:Genes, erbB-1; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D007834:Lasers; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D019076:Transgenes
D018840:HSP70 Heat-Shock Proteins