After the completion of several entire genome projects and a remarkable increase in public genetic databases in the recent years the results of post-genomic analyses can facilitate a better understanding of the genomic evolution underlying the diversity of organisms and the complexity of gene function. This influx of genomic information and resources is also beneficial to the discipline of systematic biology. In this paper, we describe a set of 6 previous and 22 new PCR/sequencing primers for RAG1, Rhodopsin and four novel nuclear markers from IRBP, EGR1, EGR2B and EGR3 that we developed through an approach making use of public genetic/genomic data mining for one of the ongoing tree of life projects aimed at understanding the evolutionary relationships of the planet's largest clade of freshwater fishes--the Cypriniformes. The primers and laboratory protocols presented here were successfully tested in 33 species comprising all cypriniform family and subfamily groups. Phylogenetic performance of each gene, as well as their implications in the investigation of the evolution of cypriniform fishes were assessed and discussed.