Cloning and characterization of a Rab11 homologue inGracilariopsis lemaneiformis

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Abstract

A small GTPase-encoding gene, homologous to Rab11s from animals, was cloned from the red alga Gracilariopsis lemaneiformis. The gene was isolated from libraries generated by suppression subtractive hybridization between female gametophytes and tetrasporophytes, designated as Gl Rab11, and found to be phase-relative, upregulated in female gametophytes. Two transcripts of Gl Rab11 were obtained by RACE (rapid amplification of cDNA ends). One was a mature mRNA and the other was an intermediate formed during mRNA processing. The former had 737 nucleotides with an open reading frame of 648 nucleotides encoding 214 amino acid residues, 83 nucleotides of 5′-UTR (untranslated-region) and 12 nucleotides of 3′-UTR. The amino acid sequence analysis of Gl Rab11 showed that it contained a Rab domain and a double cysteine motif in the C terminus. Southern blot analysis revealed that Gl Rab11 was probably a single copy gene. Two recombinant GlRab11 proteins were able to bind GTP in vitro. The identification of proteins involved in the phase development of G. lemaneiformis might shed some light on the still obscure molecular events during phase formation in red algae.

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