In model teleost fishes like the medaka and the zebrafish many genes which have been identified in genome sequencing projects await their functional characterization. Techniques for the effective generation of transgenic animals are a prerequisite for this challenging task, and, due to their transparency, fish offer the possibility to combine the use of fluorescent proteins and developmental analysis in vivo. Here we describe the application of the Ac/Ds transposon system to generate transgenic medaka reporter and gene trap lines. We determined a germline transmission rate of 30% in our experiments using constructs ranging in size from 1.8 to 6 kilobase pairs. The genomic integration site of the Ds-elements can be easily identified which is an important feature for gene trap mutagenesis experiments and similar approaches. We constructed gene trap vectors with functional elements of medaka sequences that produce in frame fusions of the endogenous sequence to EGFP. These vectors mimic endogenous expression of the trapped allele in transgenic animals and are capable to interfere with the expression of the wild type allele in the homozygous individuals.