The synthesis of different early T4 transcripts by E. coli RNA polymerase in vitro can be dissociated in two ways: (1) when T4 DNA is mechanically degraded, it selectively loses the ability to serve as a template for the transcription of delayed early T4 RNA; (2) when initiation of RNA synthesis on intact T4 DNA is limited to a very short time interval immediate early T4 RNA is synthesized at once but delayed early polynucleotide sequences appear only after a time lag. We conclude that initiation sites for T4 early RNA synthesis are closer to immediate early than to delayed early segments of the T4 template and that immediate early and delayed early segments of the T4 chromosome are interspersed. We propose that this interspersing is significant for the programming of transcription during T4 development.