Characterization of a vasa homolog in Mekong giant catfish (Pangasianodon gigas): Potential use as a germ cell marker

Abstract

For the long-term preservation of the genetic resources of endangered fish species, a combination of germ cell cryopreservation and transplantation can be an effective technique. To optimize these techniques, it is important to identify undifferentiated germ cells possessing transplantability, such as primordial germ cells, type A spermatogonia (ASGs), and oogonia. In this study, a homolog of vasa cDNA in Mekong giant catfish (MGC-vasa) (Pangasianodon gigas), which is an endangered species inhabiting the Mekong river, was cloned and characterized for use as a putative germ cell marker. Results indicate that MGC-Vasa contained all of the consensus motifs, including the arginine–glycine and arginine–glycine–glycine motifs, as well as the nine conserved motifs belonging to the DEAD-box family of proteins. Results from phylogenetic analysis indicated MGC-vasa also grouped with Vasa and was clearly distinguishable from Pl10 in other teleosts. Results from analysis of abundance of mRNA transcripts using reverse transcription-polymerase chain reaction and in situ hybridization performed on immature Mekong giant catfish testis indicated vasa was present specifically in germ cells, with large abundances of the relevant mRNA in spermatogonia and spermatocytes. Sequence similarity and the specific localization of MGC-vasa in these germ cells suggest that the sequence ascertained in this study was a vasa homolog in Mekong giant catfish. Furthermore, vasa-positive cells were detected in prepared smears of testicular cells, indicating that it may be a useful germ cell marker for enzymatically dissociated cells used for transplantation studies.