Generation and Characterization of Blood Vessel Specific EGFP Transgenic Zebrafish via To12 Transposon Mediated Enhancer Trap Screen

Cardiovasculature forms during early stages of embryonic development and enables other organs to develop, maintain and regenerate. hribalanced growth of blood vessels can give rise to numerous pathological disorders. However, the genes involved in blood vessel development remain largely elusive. Zebrafish (Banjo rerio) is an ideal vertebrate model organism for the study of developmental processes, especially for that of cardiovascular formation. 26 transgenic fish lines with blood vessel-specific EGFP expression were identified via a large scale enhancer trap screen mediated by Tol2 transposon in zebrafish. The EGFP expression in some of these lines shows different and unique patterns in different part of blood vessels. The genomic sequences flanking the Tol2 insertion sites have been successfully cloned from 22 lines via linker-mediated PCR, among which 17 sequences could be mapped to a unique location within current zebrafish genome assembly. Expression of 8 flanking genes from 9 transgenic lines was confirmed to recapitulate the expression of EGFP reporter gene in their corresponding lines via RNA whole mount in situ hybridization (ISH). Three of these genes, hhex, win, and dusp5 are known to be important for vasculargenesis. Since hhex and etsla are also expressed in hematopoietie precursors, these transgenic zebrafish should be very useful for the study of both hematopoiesis and vasculargenesis. The rest of these genes, namely zvsg1, micoll2a, arl8b (1 of 2), zgc:73355 and hecw2 (1 of 2), are either novel or functionally unknown in zebrafish. Further investigation of these fish lines and corresponding genes will give important insights of blood vessel developmental mechanisms including hemangioblast formation and differentiation, as well as genes and enhancer elements important for cardiovascular system. In addition, these transgenic fish lines could also make invaluable contributions to small molecule screen for drug discovery.