Ketocarotenoids are obtained by the action of the β-carotene ketolase, which catalyses the addition of a keto-group at the C4 position of carotenoids β-ion-rings. Most microalgae and higher plants do not posses the carotene ketolase activity and consequently do not synthesize ketocarotenoids, which are highly demanded as feed supplements and as nutraceutical for human nutrition. Here we propose the use of the unicellular microalga Chlamydomonas reinhardtii to express the Bkt (β-carotene ketolase) gene from Haematococcus pluvialis and synthesize a new ketocarotenoid that the vegetative cells of Chlamydomonas do not synthesize in the natural way. The methodology needed to successfully achieve metabolic engineering of ketocarotenoids synthesis in Chlamydomonas is described in this chapter, including the construction of a vector containing the Bkt gene, transformation of Chlamydomonas , selection of transformants, and carotenoids analysis.