Generation and Characterization of a Transgenic Zebrafish Expressing the Reverse Tetracycline Transactivator

作     者：Qilin Gu Xiaojie Yang Xiaozhen He Qing Li Zongbin Cui 

作者机构：The Key Laboratory of Aquatic Biodiversity and Conservation of Chinese Academy of SciencesInstitute of HydrobiologyChinese Academy of Sciences University of Chinese Academy of Sciences 

出 版 物：《Journal of Genetics and Genomics》 (遗传学报（英文版）)

年 卷 期：2013年第40卷第10期

页      面：523-531页

核心收录：

学科分类：0710[理学-生物学] 0908[农学-水产] 1001[医学-基础医学(可授医学、理学学位)] 09[农学] 

基　　金：supported by the grants from the National Basic Research Program of China(No.2012CB944500) the National Natural Science Foundation of China(No.31171390 to Z.Cui) 

主　　题：Zebrafish Transgene Tet-on system Reverse Tet transactivator Doxycycline 

摘      要：Conditional expression of a target gene during zebrafish development is a powerful approach to elucidate gene functions. The tetracycline-controlled systems have been successfully used in the modulation of gene expression in mammalian cells, but few lines of zebrafish carrying these systems are currently available. In this study, we had generated a stable transgenic zebrafish line that ubiquitously expressed the second-generation of reverse Tet transactivator （rtTA-M2）. Southern blotting analysis and high-throughput genome sequencing verifed that a single copy of rtTA-M2 gene had stably integrated into the zebrafish genome. After induction with doxycycline （Dox）, a strong green fluorescent protein （GFP） was seen in rtTA-transgenic eggs injected with pTRE--EGFP plasmids. The fluorescent signal gradually decreased after the withdrawal of Dox and disappeared. However, leaky expression of GFP was undetectable before Dox- induction. Additionally, transgenic embryos expressing rtTA-M2 exhibited no obvious defects in morphological phenotypes, hatching behavior and expression patterns of developmental marker genes, suggesting that rtTA-M2 had little effect on the development of transgenic zebrafish. Moreover, expressed Dickkopf-1 （DKK1） in pTRE-DKKl-injected embryos led to alterations in the expression of marker genes associated with Wnt signaling. Thus, this rtTA-transgenic zebrafish can be utilized to dissect functions of genes in a temporal manner.