Characterization of vasa and its promoter region in jundiá (Rhamdia quelen) to produce a transgenic fish line expressing green fluorescent protein under the control of the vasa promoter

Abstract

Primordial germ cells (PGCs) are the progenitors of the germ cell lineage in all living animals. In fish, PGC specification is dependent of maternal inherited factors, which are concentrated in the "germ plasm" of the embryos. In the germ plasm, vasa is one of the crucial maternal factors responsible to induce PGC differentiation. Vasa is also crucial for germ cell development and is highly expressed in early spermatogonia, including spermatogonial stem cells (SSCs). The application of transgenic techniques has been widely used to generate transgenic fish lines expressing EGFP (enhanced green fluorescent protein) under the control of vasa promoter. Such transgenic lines can be useful for several applications in biotechnology and aquaculture fields. For example, vasa::egfp fish can be useful to select (sort) SSCs and primordial germ cells for transplantation, cryopreservation and conservation studies. Thus, the aim of this project is to clone and sequence vasa and its promoter of a Brazilian important native species Jundiá (Rhamdia quelen), and then generate constructs which have egfp under control of this vasa promoter. Utilizing the Tol2 transposon system, injections of these constructs first in zebrafish (Danio rerio) embryos, and further in Jundiá embryos, will generate transgenic lines. This technique will generate the first transgenic line for germ cell lineage for a native species. Moreover, this expertise, learned from Dr. Bogerd will allow the constructions other transgenic lines, for example for other SSC markers. (AU)