Long and complex molecules of DNA are precisely organized within cells to support genome maintenance and gene expression. In addition to prokaryotic cells and eukaryotic nuclei, this principle also applies to organelles of endosymbiotic origin, which originated from bacterial ancestors and retain their own genomes. One type of endosymbiotic organelle is the chloroplast, which is present in plant and algal cells and is responsible for photosynthesis. We show that chloroplast DNA is organized by binding to membranes present inside the organelle. We propose that transcription, the initial step in the process of gene expression, plays an important role in organizing the chloroplast nucleoid into a transcriptionally active membrane-associated core and a less active periphery.