BAC Transgenic Zebrafish for Transcriptional Promoter and Enhancer Studies
With the advent of BAC recombineering techniques, transcriptional promoter and enhancer isolation studies have become much more feasible in zebrafish than in mouse given the easy access to large numbers of fertilized zebrafish eggs and offspring in general, the easy to follow ex-utero development of zebrafish, an overall less skill demand and a more cost-effective technique. Here we provide guidelines for the generation of BAC recombineering-based transgenic zebrafish for DNA transcriptional promoter and enhancer identification studies as well as protocols for their analysis, which have been successfully applied in our laboratories many times. BAC recombineering in zebrafish allows for economical functional genomics studies, for example by integrating developmental biology with comparative genomics approaches to validate potential enhancer elements of vertebrate transcription factors.
Key wordsBAC recombination technology Enhancer lacZ EGFP Zebrafish Paraffin sectioning
The authors are grateful to Serene Lee and Song Jie as well as Drs Igor Kondrychyn, V Sivakamasundari, Zhen Li, Sumantra Chatterjee, Vladimir Korzh, and especially Mathavan Sinnakaruppan for invaluable advice, patience and access to the zebrafish equipment.