| Abstract: | Degenerate primers corresponding to conserved protein kinase motifs were used to amplify potential kinase DNA fragments from a Gonyaulax polyedra Stein cDNA library using PCR. One PCR fragment, potentially encoding a CAMP-dependent protein kinase, was used as a probe to isolate a near full-length cDNA from the library. The nucleic acid sequence of the entire cDNA clone had a high homology to the catalytic subunit of cAMP-dependent protein kinase (cAPK subfamily and affiliated members. Northern blot analysis showed that the corresponding mRNA had a size (about 1.4 kb) and a relative high abundance consistent with a cAPK homologue. Southern blot analysis showed that while there are roughly 30 copies of the kinase gene per genome, the pattern of restriction fragments is inconsistent with the hypothesis of a large gene family. Phylogenetic analyses comparing the deduced amino acid sequence from the Gonyaulax cDNA with other cAPK sequences place Gonyaulax close to the slime mold Dictyostelium discoideum. This is the first phylogenetic analysis of dinoflagellates based on protein sequence, and the results are in agreement with similar analyses based on rRNA sequences. |
