Clade identification of symbiotic zooxanthellae of dominant sclerectinian coral species of intertidal pools in Hengam Island

Zooxanthellae of reef-building corals are unicellular dinoflagellates of the Symbiodinium genus, which has an important role in bleaching phenomenon. Symbiodinium and their coral hosts are sensitive to environmental stresses that include salinity, high temperatures, low temperatures, extreme light levels and turbidity. Tidal pools have harsh conditions due to lack of nutrients, food and pronounced changes in physical conditions such as pH, salinity and temperature, hence the study of symbiotic zooxanthellae on coral reefs of tidal pool seems to be necessary. Samples of five coral species that include Siderastrea savignyana, Coscinaraea columna, Anomastrea irregulariis, Cyphastrea serailia, Psammocora superficialis were collected at intertidal pool of Hengam Island in the northern Persian Gulf. Partial 28S nuclear ribosomal (nr) DNA of Symbiodinium were amplified by polymerase chain reaction (PCR) and then PCR products were analyzed by the phylogenetic analyses of the LSU DNA sequences based on PAUP and Clustal X software. The results showed that there are at least two clades of Symbiodinium from Hengam Island. Clade D was detected from 3 of the coral species whileclade C was found in 2 species only. This study showed dominance of clade D at intertidal pool in Hengam Island and the dominace of clade D might be explained by the high environmental stresses for the Persian Gulf.Key words: Persian Gulf, clade D, tides, Symbiodinium and Hengam Island

Cloning, Expression and Characterization of Zebra Fish Ferroportin in Hek 293T Cell Line

Background: Ferroportin (Fpn), a regulator of iron homeostasis is a conserved membrane protein that exports iron across the enterocytes, macrophages and hepatocytes into the blood circulation. Fpn has also critical influence on survival of microorganisms whose growth is dependent upon iron, thus preparation of Fpn is needed to study the role of iron in immunity and pathogenesis of micoorganisms.Methods: To prepare and characterize a recombinant ferroportin, total RNA was extracted from Indian zebrafish duodenum, and used to synthesize cDNA by RT-PCR. PCR product was first cloned in Topo TA vector and then subcloned into the GFP expression vector pEGFP-N1. The final resulted plasmid (pEGFP-ZFpn) was used for expression of Fpn-EGFP protein in Hek 293T cells.Results: The expression was confirmed by appearance of fluorescence in Hek 293 T cells. Recombinant Fpn was further characterized by submission of its predicted amino acid sequences to the TMHMM V2.0 prediction server (hidden Markov model), NetOGlyc 3.1 and NetNGlyc 3.1 servers. The obtained Fpn from indian zebrafish also contained eight transmembrane domains with N- and C-termini inside the cytoplasm and harboured 78 O-glycosylated amino acids.Conclusion: The recombinant Fpn from Indian zebra fish was successfully expressed in Hek 293 cell line. Although the discrepancy in two amino acids was observed in our produced Fpn and resulted in an additional O-glycosylation site, but had no effect on the topology of the protein compared to other Fpn described by other researchers. Therefore this construct can be used in future iron studies