A rod-shaped, non-endospore-forming and non-motile bacterium, strain DL-329, was isolated from the above-ground part of a plant, Androsace koso-poljanskii Ovcz. (Primulaceae), at the the State Natural Reserve 'Belogorie', Russia. On the basis of 16S rRNA gene sequence comparisons, the strain clustered with members of the genus Rathayibacter, showing the highest sequence similarity to Rathayibacter tritici (98.89 %), Rathayibacter rathayi (98.82 %) and Rathayibacter festucae (98.82 %). The DNA hybridization experiments demonstrated that strain DL-329 represents a separate genomic species. The results of comparative studies of physiological and chemotaxonomic characteristics, including cell-wall sugar patterns, polar lipid profiles, and the matrix-assisted laser desorption/ionization time-of-flight mass spectra of bacterial cells, allowed clear differentiation of VKM Ac-2121 from the recognized Rathayibacter species at the phenotypic level. Based on the data obtained, a new species, Rathayibacter oskolensis sp. nov., is proposed, with DL-329 (=VKM Ac-2121=LMG 22542) as the type strain.
Transient gene expression assays were developed to assess the function of the regulatory sequences of baculoviruses Bombyx mori nuclear polyhedrosis virus (BmNPV) and Autographa californica nuclear polyhedrosis virus (AcNPV) in insect cells of Bombyx mori and Spodoptera frugiperda, respectively. DNA sequences encoding luciferase (luc) of the firefly Photinus pyralis was successfully employed in the expression assay as a reporter gene. Recombinant plasmids were constructed containing the luc gene under control of baculovirus-specific or heterologous promoters. Cotransfection of Bombyx mori and Spodoptera frugiperda cells with recombinant plasmids carrying virus-specific promoter sequences and BmNPV and AcNPV DNA, respectively, gave rise to efficient synthesis of luciferase (Luc), while heterologous promoters induced a low level of luc expression. We found that flanking sequences of the AcNPV DNA in the transfer plasmid contained an unknown promoter conferring an efficient luc expression. The activity of this promoter was modulated by the polh promoter sequences. The assay allows one to conduct highly sensitive monitoring of the transient expression of foreign genes from the transfecting plasmids prior to construction of recombinant viruses.
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