In a series of desaturation reactions, the trienoic structures of phytoene and diapophytoene are extended to a maximum of 15 or 11 conjugated double bonds, respectively. After the cloning of several genes from bacteria and eukaryotes, the desaturation reactions were first analyzed in a heterologous host by functional genetic complementation. In addition, different desaturases were heterologously expressed and the reactions studied in vitro. This revealed that in archaea, non-photosynthetic prokaryotes and fungi the desaturases differ significantly from convergently evolved desaturases in cyanobacteria, Chlorobaculum (old name Chlorobium) species and eukaryotic photosynthetic organisms including plants. Detailed analysis of the desaturation reactions including the determination of the substrates converted by the enzymes, the intermediates and the products formed in the reactions revealed the bacterial all-trans desaturation pathway catalyzed by a single enzyme and the cyanobacterial/plant type poly-cis desaturation pathway which involves two closely related desaturases. This indicates that in the course of evolution of carotenogenesis from bacteria via cyanobacteria to plants, the simple situation of one enzyme for the entire reaction sequence from phytoene to all-trans lycopene changed to a more complex process. Three individual enzymes, newly acquired phytoene and zeta-carotene desaturases, as well as a carotene isomerase which is phylogenetically related to CrtI are involved. Only the CrtI-type enzymes seem to have the property to catalyze cis to trans conversion of carotenes.