During the last decade, methods for detecting DNA polymorphisms have proliferated at a bewildering pace. Today the investigator must choose among various types of genetic markers as well as between a variety of methods for discovering and screening polymorphisms. Polymorphisms useful for the analysis of population structure are found in both mitochondrial and nuclear genomes. Marker development may proceed along two routes: 1) discovery of species–specific markers, and 2) application of universal methods. Species-specific markers are based on sequence data from the target species or close relatives, whereas universal markers are based on the use of polymerase chain reaction (PCR) primers targeted to regions highly conserved across diverse taxa. Markers commonly employed include mitochondrial DNA polymorphisms, microsatellites, anonymous nuclear loci and known genes (both coding and noncoding regions). Methods for detecting polymorphisms range from technically simple (RFLP analysis) to more sophisticated mutation scanning methods. We review the application of these approaches to several key Antarctic species (the Patagonian toothfish Dissostichus eleginoides, the mackerel icefish Champsocephalus gunnari, and the squid Martialia hyadesi Rochebrune & Mabille, 1889) and present preliminary data on genetic polymorphisms in toothfish and icefish.