August 2015 archive

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Heterologous DNA Uptake in Cultured Symbiodinium spp. Aided by Agrobacterium tumefaciens.

PLoS One. 2015;10(7):e0132693

Authors: Ortiz-Matamoros MF, Islas-Flores T, Voigt B, Menzel D, Baluška F, Villanueva MA

Abstract
Plant-targeted pCB302 plasmids containing sequences encoding gfp fusions with a microtubule-binding domain; gfp with the fimbrin actin-binding domain 2; and gfp with AtRACK1C from Arabidopsis thaliana, all harbored in Agrobacterium tumefaciens, were used to assay heterologous expression on three different clades of the photosynthetic dinoflagellate, Symbiodinium. Accessibility to the resistant cell wall and through the plasma membrane of these dinoflagellates was gained after brief but vigorous shaking in the presence of glass beads and polyethylene glycol. A resistance gene to the herbicide Basta allowed appropriate selection of the cells expressing the hybrid proteins, which showed a characteristic green fluorescence, although they appeared to lose their photosynthetic pigments and did not further divide. Cell GFP expression frequency measured as green fluorescence emission yielded 839 per every 106 cells for Symbiodinium kawagutii, followed by 640 and 460 per every 106 cells for Symbiodinium microadriaticum and Symbiodinium sp. Mf11, respectively. Genomic PCR with specific primers amplified the AtRACK1C and gfp sequences after selection in all clades, thus revealing their presence in the cells. RT-PCR from RNA of S. kawagutii co-incubated with A. tumefaciens harboring each of the three vectors with their respective constructs, amplified products corresponding to the heterologous gfp sequence while no products were obtained from three distinct negative controls. The reported procedure shows that mild abrasion followed by co-incubation with A. tumefaciens harboring heterologous plasmids with CaMV35S and nos promoters can lead to expression of the encoded proteins into the Symbiodinium cells in culture. Despite the obvious drawbacks of the procedure, this is an important first step towards a stable transformation of Symbiodinium.

PMID: 26167858 [PubMed – in process]

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Single-cell transcriptomics using spliced leader PCR: Evidence for multiple losses of photosynthesis in polykrikoid dinoflagellates.
BMC Genomics. 2015;16:528
Authors: Gavelis GS, White RA, Suttle…

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The large mitochondrial genome of Symbiodinium minutum reveals conserved non-coding sequences between dinoflagellates and apicomplexans.
Genome Biol Evol. 2015 Jul 20;
Authors: Shoguchi E, Shinzato …

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Genome sequence of the Roseovarius mucosus type strain (DSM 17069(T)), a bacteriochlorophyll a-containing representative of the marine Roseobacter group isolated from the dinoflagellate Alexandrium ostenfeldii.

Stand Genomic Sci. 2015;10:17

Authors: Riedel T, Spring S, Fiebig A, Scheuner C, Petersen J, Göker M, Klenk HP

Abstract
Roseovarius mucosus Biebl et al. 2005 is a bacteriochlorophyll a-producing representative of the marine Roseobacter group within the alphaproteobacterial family Rhodobacteraceae, which was isolated from the dinoflagellate Alexandrium ostenfeldii. The marine Roseobacter group was found to be abundant in the ocean and plays an important role for global and biogeochemical processes. Here we describe the features of the R. mucosus strain DFL-24(T) together with its genome sequence and annotation generated from a culture of DSM 17069(T). The 4,247,724 bp containing genome sequence encodes 4,194 protein-coding genes and 57 RNA genes. In addition to the presence of four plasmids, genome analysis revealed the presence of genes associated with host colonization, DMSP utilization, cytotoxins, and quorum sensing that could play a role in the interrelationship of R. mucosus with the dinoflagellate A. ostenfeldii and other marine organisms. Furthermore, the genome encodes genes associated with mixotrophic growth, where both reduced inorganic compounds for lithotrophic growth and a photoheterotrophic lifestyle using light as additional energy source could be used.

PMID: 26203330 [PubMed]