Cloning and sequence analysis of a novel member of the rab gene family from Euplotes octocarinatus

Rab proteins belong to a subfamily of small GTP-binding proteins of the Ras superfamily,which play an important role in intracellular vesicular traffic.In this study,a rab gene was obtained from Euplotes octocarinatus by polymerase chain reaction(PCR) and RT-PCR.The rab gene from macronucleic DNA was 884 bp in length,including non-coding regions and telomeric sequences at both ends.The rab gene from micronuclear DNA(723 bp),lacking of internal eliminated sequences,was identical to rab gene from macronuclear DNA.RT-PCR showed that the opening reading frame of the rab gene was 663 bp long.The rab gene from macronuclear DNA contained an intron of 60 bp at the position from 153 bp to 212 bp of macronuclear DNA.The rab gene had two in-frame TGAs encoding for cysteine in Euplotes octocarinatus.The rab gene used TAG as stop codon,which was the first report in Euplotes octocarinatus.The result of BLAST in NCBI demonstrates that the Rab shares a homology of 49%~52% at the amino acid level with Rab1 proteins from a number of other eukaryote,which suggesting that the Rab is a Rab1 homolog.The rab gene was therefore designated Eo-rab-1N(GenBank accession number: DQ105562).The evolution of Eo-rab-1N was analyzed using phylogenetic tree of amino acids sequences of Rab1 obtained from GenBank.