PUBLICATION
One-, two-, and three-color whole-mount in situ hybridization to Drosophila embryos
- Authors
- Hauptmann, G.
- ID
- ZDB-PUB-010514-5
- Date
- 2001
- Source
- Methods (San Diego, Calif.) 23(4): 359-372 (Review)
- Registered Authors
- Hauptmann, Giselbert
- Keywords
- none
- MeSH Terms
-
- Alkaline Phosphatase/metabolism
- Animals
- Biotin/pharmacology
- Digoxigenin/pharmacology
- Drosophila
- Embryo, Nonmammalian/metabolism*
- In Situ Hybridization/methods*
- Molecular Probes/pharmacology
- RNA, Messenger/metabolism
- beta-Galactosidase/metabolism
- PubMed
- 11316437 Full text @ Methods
Citation
Hauptmann, G. (2001) One-, two-, and three-color whole-mount in situ hybridization to Drosophila embryos. Methods (San Diego, Calif.). 23(4):359-372.
Abstract
This article contains detailed protocols for the localization of mRNA transcripts within whole Drosophila embryos. The procedures are based on the use of digoxigenin-, fluorescein-, and biotin-labeled antisense RNA probes for nonradioactive detection of transcripts. The labels are visualized in situ by differently colored water-insoluble precipitates using alkaline phosphatase- or beta-galactosidase-based immunoassays. First, a basic method is described that allows detection of transcript distribution(s) of one or more genes using the same color precipitate. Second, a sequential alkaline phosphatase detection method is presented that permits the visualization of two or three independent transcript patterns in multiple colors in the same embryo. Third, a shortened two-color in situ hybridization protocol is provided that employs a combination of beta-galactosidase and alkaline phosphatase colorimetric reactions for differential detection. The two-color in situ hybridization methods work equally well in Drosophila and zebrafish embryos and may therefore also be adaptable to other species.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping