Eukaryotic cells arose through endosymbiotic uptake of free-living bacteria followed by massive gene transfer from the genome of the endosymbiont to the host nuclear genome. Because this gene transfer took place over a time scale of hundreds of millions of years, direct observation and analysis of primary transfer events has remained difficult. Hence, very little is known about the evolutionary frequency of gene transfer events, the size of transferred genome fragments, the molecular mechanisms of the transfer process, or the environmental conditions favoring its occurrence. We describe here a genetic system based on transgenic chloroplasts carrying a nuclear selectable marker gene that allows the efficient selection of plants with a nuclear genome that carries pieces transferred from the chloroplast genome. We can select such gene transfer events from a surprisingly small population of plant cells, indicating that the escape of genetic material from the chloroplast to the nuclear genome occurs much more frequently than generally believed and thus may contribute significantly to intraspecific and intraorganismic genetic variation.T he evolutionary origin of eukaryotic cells is characterized by the endosymbiotic uptake of bacteria and their gradual conversion into the DNA-containing cell organelles, mitochondria, and plastids (chloroplasts) (1-3). Genetically, the evolutionary optimization of endosymbiosis was accompanied by the loss of dispensable and redundant genetic information and the large-scale translocation of genetic information from the endosymbiont to the host genome (4-6). Consequently, contemporary organellar genomes are greatly reduced and contain only a small proportion of the genes that their free-living ancestors had possessed. By using molecular methods, the origins of organelles have been traced back to specific taxa of eubacteria: Whereas cyanobacteria were identified as presumptive ancestors of plastids, ␣-proteobacteria are related most closely to mitochondria (1).Interspecific variation in the gene content of organellar genomes (7-11) suggests that gene transfer from organelles to the nucleus is an ongoing process. Moreover, pieces of chloroplast and mitochondrial DNA are often found in nuclear genomes (12-18) and commonly referred to as promiscuous DNA. These sequences lack any apparent function but may provide the raw material for converting organellar genes into functional nuclear genes, the products of which are reimported into the organelle, then allowing for subsequent loss of the genes from the organellar genome (10). In addition, promiscuous DNA of mitochondrial origin has been implicated recently in DNA repair in yeast by patching broken chromosomes (19,20).In the present study we developed an experimental system suitable for selecting and analyzing DNA transfer events from the chloroplast genome to the nuclear genome. We find that DNA escape out of the chloroplast and integration into the nuclear genome occurs much more frequently than generally believed and thus provides a mechan...