We have constructed hybrid plasmids pColD and pIbpA in which transcription of reporter genes luxCDABE from Photohabdus luminescens occurred from induced promoters: SOS-promoter of gene cdaA (pColD-CA23) and heat shock gene ibpA (E. coli MG1655 genome), respectively. Main parameters were measured: threshold concentration of an inducer and maximal response and minimal time of response for E. coli lux-biosensors bearing hybrid plasmids pColD and pRecA (SOS-response); pIbpA and pGrpe (heat shock) and pKatG and pSoxS (oxidative stress). Mitomicyn C, cis-diaminedichloroplatinum (SOS-response), ethanol and pentachlorophenol (heat shock), and hydrogen peroxide and methyl viologen (oxidative stress) were used as luminescence inducers. It was shown that the SOS-biosensor pColD is superior in main characteristics to the earlier constructed biosensor pRecA, and the heat biosensor pIbpA is more efficient than the earlier constructed pGrpE. Lux-biosensors (pKatG) and (pSoxS) are high sensitive to hydrogen peroxide and methyl viologen, respectively.

Citrobacter freundii cells produce l -methionine γ-lyase when grown on a medium containing l -methionine. The nucleotide sequence of the hybrid plasmid with a C. freundii EcoRI insert of about 3.0 kbp contained two open reading frames, consisting of 1,194 nucleotides and 1,296 nucleotides, respectively. The first one (denoted megL ) encoded l -methionine γ-lyase. The enzyme was overexpressed in Escherichia coli and purified. The second frame encoded a protein belonging to the family of permeases. Regions of high sequence identity with the 3′-terminal part of the C. freundii megL gene located in the same regions of Salmonella enterica serovar Typhimurium, Shigella flexneri , E. coli , and Citrobacter rodentium genomes were found.

This is a review of relevant Raman spectroscopy (RS) techniques and their use in structural biology, biophysics, cells, and tissues imaging towards development of various medical diagnostic tools, drug design, and other medical applications. Classical and contemporary structural studies of different water-soluble and membrane proteins, DNA, RNA, and their interactions and behavior in different systems were analyzed in terms of applicability of RS techniques and their complementarity to other corresponding methods. We show that RS is a powerful method that links the fundamental structural biology and its medical applications in cancer, cardiovascular, neurodegenerative, atherosclerotic, and other diseases. In particular, the key roles of RS in modern technologies of structure-based drug design are the detection and imaging of membrane protein microcrystals with the help of coherent anti-Stokes Raman scattering (CARS), which would help to further the development of protein structural crystallography and would result in a number of novel high-resolution structures of membrane proteins—drug targets; and, structural studies of photoactive membrane proteins (rhodopsins, photoreceptors, etc.) for the development of new optogenetic tools. Physical background and biomedical applications of spontaneous, stimulated, resonant, and surface- and tip-enhanced RS are also discussed. All of these techniques have been extensively developed during recent several decades. A number of interesting applications of CARS, resonant, and surface-enhanced Raman spectroscopy methods are also discussed.

Gene-specific silencing refers to a phenomenon in which expression of an individual gene can be specifically repressed by different mechanisms on the levels of transcription, RNA splicing, transport, degradation in nuclei or cytoplasm, or blocking of translation. In different species gene-specific silencing was observed by expression or injections of antiparallel double-stranded RNA formed by a fragment of mRNA and antisense RNA. Here we show a potent and specific gene silencing in bacteria by expression of RNA, that is complementary in a parallel orientation to Escherichia coli lon mRNA. Moreover, the expression of parallel RNA is more effective at producing interference than expression of antisense RNA corresponding to the same mRNA region. Both effects of interference mediated either by parallel RNA or antiparallel RNA gradually decrease up to the 40th generation. Together with in vitro nuclease protection studies these results indicate that a parallel RNA duplex might be formed in vivo and both types of duplexes, antiparallel or parallel, can induce gene-specific silencing by similar mechanisms.