269 José-Luis Barredo (ed.), Microbial Carotenoids from Bacteria and Microalgae: Methods and Protocols, Methods in Molecular Biology, vol. 892, DOI 10.1007/978-1-61779-879-5_16, © Springer Science+Business Media, LLC 2012 Chapter 16 DNA Fingerprinting Intron-Sizing Method to Accomplish a Specific, Rapid, and Sensitive Identification of Carotenogenic Dunaliella Species Jorge Olmos-Soto, J. Paniagua-Michel, Rosalía Contreras, and Leonel Ochoa Abstract Dunaliella salina has become the most important microorganism for the production of β-carotene around the world. Natural carotenoids are a source of active metabolites utilized in different areas of food nutri- tion and pharmaceuticals, both in humans and also in animals. Identification of Dunaliella species from natural environments or certified culture collections is not precise and it is time consuming. However, accurate identification is extremely important because a slight difference in Dunaliella species generates great differences in carotenoids production. Here, we describe an intron-sizing method to make a rapid and precise identification for each of the most important carotenogenic species, showing that each hyper- producer species has an exclusive 18S rDNA fingerprint profile. Key words: Dunaliella, Molecular identification, Specific oligonucleotides, 18S rDNA, Fingerprint Dunaliella was originally described by Teodoresco in 1905. Since then, taxonomic studies among Dunaliella have identified several new species. However, even today, differentiation among halo- philic and carotenogenic Dunaliella species in both green and red stages is difficult and time consuming. In addition, some strains and species in culture collections are misnamed and have given rise to unnecessary strains and species names (1). This confusion of strains and species names makes comparison of results by different authors difficult (2). Dunaliella salina and D. salina/bardawil are the only reported β-carotene hyperproducer species of the genus that grows in salt-saturated lagoons (3). However, a controversy still exists about identification of D. salina/ bardawil as a different 1. Introduction