Synthesis of a new substrate for detection of lacZ gene expression in live Drosophila embryos.
Escherichia coli lacZ, which encodes beta-galactosidase, has become a widely used reporter gene to study the developmental regulation of gene expression in a variety of organisms. To detect the presence of the beta-galactosidase, the sample must be fixed and appropriately stained. This sort of analysis yields rather crude estimates of the spatial-temporal changes in gene expression patterns. In addition, one cannot recover interesting specimens for propagation. A novel fluorogenic beta-galactosidase substrate for use in live Drosophila melanogaster embryos has been designed and synthesized. This compound provides a means to determine gene dosage in live embryos so that one can unambiguously determine the genotype of a living embryo. This will be useful for detailed analysis of cellular and morphogenetic behavior changes in live embryos that are homozygous for embryonic lethal mutations. In the course of testing this compound, a new beta-galactoside hydrolytic activity, different from the previously identified beta-galactosidase, has been discovered to reside in macrophages and the intervitelline space.
Minden
J S
JS
Department of Biological Sciences, Carnegie Mellon University, Pittsburgh, PA 15213, USA.
D000818:Animals; D004331:Drosophila melanogaster / Q000196:embryology / Q000235:genetics*; D005456:Fluorescent Dyes / Q000138:chemical synthesis; D005697:Galactosides / Q000138:chemical synthesis; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D005838:Genotype; D007763:Lac Operon; D010078:Oxazines / Q000138:chemical synthesis; D011092:Polyethylene Glycols; D013379:Substrate Specificity; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D005456:Fluorescent Dyes; D005697:Galactosides; D010078:Oxazines; C098903:resorufin galactoside bis(aminopropyl)polyethylene glycol 1900; D011092:Polyethylene Glycols; D001616:beta-Galactosidase