We have compared the strength of a trout protamine gene promoter with that of the mouse beta major-globin gene by analysing the relative levels of run-off transcripts produced in a single mammalian in vitro transcription reaction. When the promoters are introduced on separate recombinant plasmids, the protamine transcripts are synthesised with much greater efficiency than those originating from the globin cap site. This enhanced transcription of the protamine gene is again observed when the promoters are applied as separate DNA fragments derived from the same recombinant plasmid. However, when the promoters are linked on a DNA fragment that includes 7 kb of DNA separating the initiation sites, then there is a marked reduction in the protamine signal relative to the globin. Deletion of a region of this fragment that contains the sequences flanking the globin gene at positions -335 to -1400 restores the enhanced protamine gene expression to the levels observed when the promoters are carried on separate DNA fragments.