Penaeus stylirostris densovirus (PstDNV) genome contains three open reading frames (ORFs), left, middle, and right, which encode a non-structural (NS) protein, an unknown protein, and a capsid protein (CP), respectively. Transcription mapping revealed that P2, P11 and P61 promoters transcribe the left, middle and right ORFs. NS transcript uses the D1/A1 donor/acceptor sites for splicing and has two alternate transcription termination sites (TTS) that were different from the previously predicted TTS. The transcription initiation site (TIS) and the TTS for the middle and the right ORFs conform to predicted sites. PstDNV transcript quantification in infected shrimp revealed that the NS and CP transcripts were expressed at an equivalent level and significantly higher than the middle ORF transcript. In vitro assay showed that P2 had the highest promoter activity followed by P11 and P61. Transcription mapping data provided new insights into PstDNV gene expression strategy.