The significance of polyunsaturated fatty acids (PUFA) to humans lies in their biological activity, as precursors for groups of nutritionally important compounds and as essential cellular components. They are also crucial in animals for the proper development of retinal and nervous tissue. In this work, Antarctic sea samples were used to isolate bacterial cultures producing PUFA. Three bacterial strains were isolated from three different samples and they were named as BRI 1, BRI 7 and BRI 28. Percentage of eicosapentaenoic acid (EPA) was 42.7 and 42.6 of total fatty acids in BRI 7 and 28 respectively. Whereas, docosahexaenoic acid (DHA) (1.4%) was detected only in BRI 28. BRI 7 and 28 showed considerable similarities in their fatty acid profiles, hence these three isolates were further used for random amplification of polymorphic DNA (RAPD) analysis using nine random primers to confirm their genetic relatedness. In contrast to fatty acid profiles, maximum polymorphism (58%) was obtained between BRI 7 and 28, while maximum monomorphism (83.63%) was observed between BRI 1 and 7. Our results indicate that, although the conservation of PUFA genes may point to their potential utilization as phylogenetic markers in culture-independent ecological studies, the biosynthetic diversity of bacterial PUFA production has yet to be fully described.