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Chloroplast DNA from Euglena gracilis was separated in an alkaline CsC1 density gradient into a heavy and a light component. The heavy and the light DNA components each contain 18 percent self-complementary nucleotide sequences. Equal masses of the heavy and the light components do not contain equal molar complementary nucleotide sequences. The light DNA component contains sequences complementary to the majority of the chloroplast DNA transcripts. The heavy DNA component contains sequences complementary to chloroplast ribosomal RNA.

The fraction of the chloroplast DNA transcribed in the single celled alga Euglena has been determined by RNA-DNA hybridization. A vast excess of total cell RNA from cells which were rapidly dividing in the light was hybridized in liquid to [125I] — chloroplast DNA, and the resulting duplexes separated on hydroxyapatite columns. The contribution of DNA-DNA duplex formation was determined separately and was used to calculate that portion of the duplex which was actually a RNA-DNA hybrid. Sixteen percent of the single stranded chloroplast DNA forms a duplex with this RNA suggesting that 32 percent of the double stranded DNA molecule is being transcribed into RNA under these conditions of cell growth.

Transcription of Euglena chloroplast DNA was visualized by autoradiography of hybrids formed between electrophoretically separated EcoRI endonuclease fragments and [32P]-pulse labeled RNA prepared from cells at different stages of chloroplast development. The RNA was hybridized to Southern imprints of EcoRI chloroplast DNA digests. The EcoRI chloroplast DNA fragments were categorized by their ability to hybridize the different pulse labeled RNA's. Some fragments were transcribed: 1. both in the dark and throughout chloroplast development, 2. predominantly in the light, 3. in the dark and for only a brief period of time after transfer of the cells to the light. Other fragments were probably not transcribed.