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Zebrafish enhancer detection (ZED) vector: A new tool to facilitate transgenesis and the functional analysis of cis ‐regulatory regions in zebrafish

2009; Wiley; Volume: 238; Issue: 9 Linguagem: Inglês

10.1002/dvdy.22051

1097-0177

José Bessa, Juan J. Tena, Elisa de la Calle‐Mustienes, Ana Fernández‐Miñán, Silvia Naranjo, Almudena Fernández, Lluı́s Montoliu, Altuna Akalin, Boris Lenhard, Fernando Casares, José Luis Gómez-Skármeta,

Animal Genetics and Reproduction

Abstract The identification and characterization of the regulatory activity of genomic sequences is crucial for understanding how the information contained in genomes is translated into cellular function. The cis ‐regulatory sequences control when, where, and how much genes are transcribed and can activate (enhancers) or repress (silencers) gene expression. Here, we describe a novel Tol2 transposon‐based vector for assessing enhancer activity in the zebrafish ( Danio rerio ). This Zebrafish Enhancer Detector (ZED) vector harbors several key improvements, among them a sensitive and specific minimal promoter chosen for optimal enhancer activity detection, insulator sequences to shield the minimal promoter from position effects, and a positive control for transgenesis. Additionally, we demonstrate that highly conserved noncoding sequences homologous between humans and zebrafish largely with enhancer activity largely retain their tissue‐specific enhancer activity during vertebrate evolution. More strikingly, insulator sequences from mouse and chicken, but not conserved in zebrafish, maintain their insulator capacity when tested in this model. Developmental Dynamics 238:2409–2417, 2009. © 2009 Wiley‐Liss, Inc.