Abstract One of the most common transitions observed in actinopterygian fishes were from marine to freshwater environments. This phenomenon could suggest that osmoregulatory mechanisms in this group are particularly adaptable to salinity changes. The silversides of genus Odontesthes are not the exception within Atherinomorpha, with several invasions from marine to freshwater environments reported for this group. In this work, a de-novo transcriptome assembly of Odonthestes argentinensis gill tissue was performed, by means of paired-end sequencing in an Illumina HiSeq 2000 platform. Analyses of the six transcriptomes obtained result in 12,219 genes, recovered from a constructed reference transcriptome. Functional classification performed with DAVID retrieved information for 11,412 of the 12,219 annotated genes, with twenty-four GO terms and 10 KEGG pathways significantly enriched. Additional analyses performed with PANTHER suggested that “Ubiquitin proteasome pathway” was significantly overrepresented and the “Cadherin signaling pathway” was subrepresented against the DAVID database for D. rerio. Taking into account the 12,219 annotated genes, 12,189 of them were found expressed by Kallisto in at least one individual, with 42 GO terms significantly enriched compared against the gene list obtained from the reference transcriptome, as well as 5 pathways from the KEGG database and 2 from PANTHER; the last gene list analysis mentioned also found 12 Protein Classes significantly overrepresented. Considering SSR markers found in specific genes, a total of 1216 primer pairs were designed that could be used in further studies in O. argentinensis and related species, and in conjunction with the 10,784 potential SNPs were identified in annotated genes, results in set of molecular markers that could be used in future mapping or population genetic studies. Taken as a whole, the results obtained in this work will be helpful to study how individuals of genus Odontesthes deals with environmental salinity changes, as well as in studies of population structure and demographic history in this species and related taxa.