Promoter-active fragments of Synechococcus PCC7942 were isolated by transcriptional gene fusion to the promoterless beta-glucuronidase (GUS) gene of E. coli, which was used as a reporter gene. Several of the isolated promoter-active fragments expressed GUS activity in Synechococcus comparable to that of the lambda P-R promoter. Only 10% of the isolated promoter-active fragments also functioned in E, coli. The transcription initiation sites of the two promoter-active fragments, D13 and E3, were identified. The major transcription initiation sites of D13 and E3 in Synechococcus were located within the nucleotides TTTG and TTG respectively, which were identical to those corresponding to E. coli. The inferred -10 and -35 regions of D13 were TAAACT and TTGTAG respectively, which conformed to the E. coli sigma(70) promoter. Immediately upstream of the E3 transcription initiation sites was the tRNA(pro) (GGG) gene, which contained two regions exhibiting strong homology to the major promoter elements in eukaryotic tRNA genes, but did not contain the E. coli promoter element. Thus, the tRNA(pro) gene can act as a promoter.