We have used electron microscopic RNA loop mapping to determine, to within 200 nucleotides, the chromosome coordinates of the 5′ and 3′ ends of adenovirus type 2 transcripts isolated from the cytoplasm of productively infected human KB cells. The major blocks of transcription and many of the individual transcripts have discrete lengths and well defined 5′ and 3′ termini. Early cytoplasmic RNA is transcribed from four separate regions of the adenovirus genome, in map intervals 1.3-11.1, 62.4-67.9, 78.6-86.1 and 91.5-96.8. The most frequent late RNA loop occurred between coordinates 51.9 and 62.2. A late transcript complementary to interval 86.3-91.5 formed a characteristic R loop with a long branch of displaced RNA from coordinate 89.8-91.5, possibly a result of secondary structure near coordinate 89.8. Other late cytoplasmic RNA loops were located between coordinates 11.1 and 14.9, and in the intervals from 30-52 and 68-83. Transcripts from several regions of the chromosome had variable lengths but common 5′ or 3′ termini. Few R loops were seen between map coordinates 15 and 30. Switches between template strands were observed at map positions 11.2, 62.4 and 91.5. Many of the R loops have been correlated with gene products mapped previously by translation of RNA complementary to different portions of the chromosome. © 1977.