Identification and transcription of transfer RNA genes in dinoflagellate plastid minicircles

The dinoflagellate chloroplast genome is unique in that the genes are found on small circular DNA molecules carrying from one to three genes. In addition, only 14 of the typical chloroplast-located genes have so far been discovered on minicircles, while a number have been transferred to the nucleus. We have sequenced four new minicircles from the dinoflagellate Heterocapsa triquetra, three of which carry a single protein-coding gene (psbD, psbE, petD) and one that appears to be an "empty" circle. Using the tRNA prediction programs ARAGORN and tRNAscan-SE, tRNA-Met was found in the petD circle immediately downstream of the end of petD, while tRNA-Trp and tRNA-Pro were detected in the psbE and petD circles as well as in several chimeric circles of H. triquetra and the psbA minicircles of Heterocapsa pygmaea. RT-PCR showed that the tRNAs were co-transcribed with the protein-coding genes that preceded them, and cleaved from the precursor before a poly(U) tail was added to the mRNA.