Abstract Prodynorphin gene transcripts have been characterized in human tissues with cRNA probes covering parts of the non-coding exon 1 and the main coding exon 4 using Northern blot and RNase protection experiments. A 2.8-kb signal was observed in human brain RNA with both the exon 1 and exon 4 probes. An RNase protection assay with the exon 1 probe, performed to map the 5′ end of this transcript, produced protected fragments in the range of 0.11 to 0.15 kb,indicating that exon 1 is 1.2 kb shorter than previously proposed. 5′-RACE-PCR and sequencing of amplified cDNA fragments confirmed this assignment. In adrenal gland, testis and the human small cell lung carcinoma cell line, U1690, several prodynorphin-like mRNAs structurally different from the brain prodynorphin mRNA species were observed.