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#24312414   2013/12/06 Save this To Up

Gene expression system in green sulfur bacteria by conjugative plasmid transfer.

Gene transfer and expression systems in green sulfur bacteria were established by bacterial conjugation with Escherichia coli. Conjugative plasmid transfer from E. coli S17-1 to a thermophilic green sulfur bacterium, Chlorobaculum tepidum (formerly Chlorobium tepidum) WT2321, was executed with RSF1010-derivative broad-host-range plasmids, named pDSK5191 and pDSK5192, that confer erythromycin and streptomycin/spectinomycin resistance, respectively. The transconjugants harboring these plasmids were reproducibly obtained at a frequency of approximately 10(-5) by selection with erythromycin and a combination of streptomycin and spectinomycin, respectively. These plasmids were stably maintained in C. tepidum cells in the presence of these antibiotics. The plasmid transfer to another mesophilic green sulfur bacterium, C. limnaeum (formerly Chlorobium phaeobacteroides) RK-j-1, was also achieved with pDSK5192. The expression plasmid based on pDSK5191 was constructed by incorporating the upstream and downstream regions of the pscAB gene cluster on the C. tepidum genome, since these regions were considered to include a constitutive promoter and a ρ-independent terminator, respectively. Growth defections of the ∆cycA and ∆soxB mutants were completely rescued after introduction of pDSK5191-cycA and -soxB that were designed to express their complementary genes. On the other hand, pDSK5191-6xhis-pscAB, which incorporated the gene cluster of 6xhis-pscA and pscB, produced approximately four times more of the photosynthetic reaction center complex with His-tagged PscA as compared with that expressed in the genome by the conventional natural transformation method. This expression system, based on conjugative plasmid, would be applicable to general molecular biological studies of green sulfur bacteria.

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#20569108   2010/08/16 Save this To Up

Hyperthermia-induced apoptosis in Tca8113 cells is inhibited by heat shock protein 27 through blocking phospholipid scramblase 3 phosphorylation.

Hyperthermia induces tumour cell apoptosis through the mitochondrial apoptotic pathway; however, the signal transduction mechanism underlying this process still needs to be fully elucidated. Phospholipid scramblase 3 (PLS3), a target of protein kinase C-delta (PKC-delta), resides in mitochondria and plays pivotal roles in regulating apoptotic response. Activated PLS3 facilitates cardiolipin (CL) translocation from the mitochondrial inner membrane to the outer leaflet of the mitochondrial outer membrane and triggers apoptosis.

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Heat Shock Protein 27, hu Apoptosis Phospho-Specifi Rabbit Anti-APIP Apaf1 In Rabbit Anti-APIP Apaf1 In Rabbit Anti-APIP Apaf1 In Rabbit Anti-TNIP2 ABIN2 T Rabbit Anti-Azurocidin Ca Rabbit Anti-Azurocidin Ca Rabbit Anti-Azurocidin Ca Rabbit Anti-Cell death in Rabbit Anti-G protein alp Human Heat shock proteins

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#15554903   2005/04/01 Save this To Up

TraA and its N-terminal relaxase domain of the Gram-positive plasmid pIP501 show specific oriT binding and behave as dimers in solution.

TraA is the DNA relaxase encoded by the broad-host-range Grampositive plasmid pIP501. It is the second relaxase to be characterized from plasmids originating from Gram-positive organisms. Full-length TraA (654 amino acids) and the N-terminal domain (246 amino acids), termed TraAN246, were expressed as 6xHis-tagged fusions and purified. Small-angle X-ray scattering and chemical cross-linking proved that TraAN246 and TraA form dimers in solution. Both proteins revealed oriTpIP501 (origin of transfer of pIP501) cleavage activity on supercoiled plasmid DNA in vitro. oriT binding was demonstrated by electrophoretic mobility shift assays. Radiolabelled oligonucleotides covering different parts of oriTpIP501 were subjected to binding with TraA and TraAN246. The KD of the protein-DNA complex encompassing the inverted repeat, the nick site and an additional 7 bases was found to be 55 nM for TraA and 26 nM for TraAN246. The unfolding of both protein constructs was monitored by measuring the change in the CD signal at 220 nm upon temperature change. The unfolding transition of both proteins occurred at approx. 42 degrees C. CD spectra measured at 20 degrees C showed 30% a-helix and 13% b-sheet for TraA, and 27% alpha-helix and 18% beta-sheet content for the truncated protein. Upon DNA binding, an enhanced secondary structure content and increased thermal stability were observed for the TraAN246 protein, suggesting an induced-fit mechanism for the formation of the specific relaxase-oriT complex.

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Bovine Androstenedione,AS Rabbit Anti-Rat Androgen Multiple organ tumor tiss EpiQuik MBD2 Binding Acti  EpiQuik MBD2 Binding Ac BCIP INT Solution BCIP INT Solution BCIP INT Solution Astra Blue Solution Astra Blue Solution Gram's Decolorizer Solut Gram's Decolorizer Solut

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