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Chloroplast genomes of land plants and algae contain generally between 100 and 150 genes. These genes are involved in plastid gene expression and photosynthesis and in various other tasks. The function of some chloroplast genes is still unknown and some of them appear to be essential for growth and survival. Repressible and reversible expression systems are highly desirable for functional and biochemical characterization of these genes. We have developed a genetic tool that allows one to regulate the expression of any coding sequence in the chloroplast genome of the unicellular alga Chlamydomonas reinhardtii. Our system is based on vitamin-regulated expression of the nucleus-encoded chloroplast Nac2 protein, which is specifically required for the expression of any plastid gene fused to the psbD 5′UTR. With this approach, expression of the Nac2 gene in the nucleus and, in turn, that of the chosen …