title: The use of whole-mount in situ hybridization to illustrate gene expression regulation.
abstract: In situ hybridization is a widely used technique for studying gene expression. Here, we describe two experiments addressed to postgraduate genetics students in which the effect of transcription factors on gene expression is analyzed in Drosophila embryos of different genotypes by whole-mount in situ hybridization. In one of the experiments, students analyzed the repressive effect of Snail over rhomboid expression using reporter lines containing different constructs of the rhomboid neuroectodermal enhancer fused to the lacZ gene. In the second experiment, the epistatic relationship between the cabut and decapentaplegic genes was analyzed. These simple experiments allowed students to (1) understand the role of transcription factors and cis-regulatory elements over gene expression regulation and (2) practice a widespread laboratory technique, in situ hybridization with nonradioactive labeled probes, to detect gene expression patterns. These experiments required 12 hr and were organized into four daily sessions that included the discussion of the results with students. Examples of the results obtained and their relevance are shown and discussed herein. The methods described in these laboratory exercises can be easily adapted to model organisms other than Drosophila.
authors: Department of Genetics, Faculty of Biology, Universitat de València, Valencia, Spain; INCLIVA Health Research Institute, Valencia, Spain.
authors: Beatriz
authors: B
authors: Llamusí
authors: Verónica
authors: V
authors: Muñoz-Soriano
authors: Nuria
authors: N
authors: Paricio
authors: Rubén
authors: R
authors: Artero
mesh_terms: D000818:Animals; D004330:Drosophila / Q000196:embryology / Q000235:genetics*; D029721:Drosophila Proteins / Q000235:genetics; D004500:Education, Graduate / Q000379:methods*; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental; D017403:In Situ Hybridization / Q000379:methods*; D008565:Membrane Proteins / Q000235:genetics; D008967:Molecular Biology / Q000193:education* / Q000379:methods; D015203:Reproducibility of Results; D000071250:Snail Family Transcription Factors; D013663:Teaching / Q000379:methods; D014157:Transcription Factors / Q000235:genetics
chemical_list: C506602:Cbt protein, Drosophila; D029721:Drosophila Proteins; D008565:Membrane Proteins; C081481:Rho protein, Drosophila; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; C064196:dpp protein, Drosophila; C000604451:sna protein, Drosophila
keywords: Drosophila; cabut; decapentaplegic; enhancer; rhomboid; signaling pathway