SummaryIn Escherichia coli, the transcriptional activity of many promoters is strongly dependent on the negative superhelical density of chromosomal DNA. This, in turn, varies with the growth phase, and is correlated with the overall activity of DNA gyrase, the major topoisomerase involved in the elevation of negative superhelicity. The DNA architectural protein FIS is a regulator of the metabolic reorganization of the cell during early exponential growth phase. We have previously shown that FIS modulates the superhelical density of plasmid DNA in vivo, and on binding reshapes the supercoiled DNA in vitro. Here, we show that, in addition, FIS represses the gyrA and gyrB promoters and reduces DNA gyrase activity. Our results indicate that FIS determines DNA topology both by regulation of topoisomerase activity and, as previously inferred, by directly reshaping DNA. We propose that FIS is involved in coupling cellular physiology to the topology of the bacterial chromosome.

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We present novel multiplex PCR methods for rapid and reliable screening of genetically modified organisms (GMOs). New designed PCR primers targeting four frequently used GMO specific sequences permitted identification of new DNA markers, in particular 141 bp fragment of cauliflower mosaic virus (CaMV) 35S promoter, 224 bp fragment of Agrobacterium tumefaciens nopaline synthase (NOS) terminator, 256 bp fragment of 5-enolppyruvylshikimate-phosphate synthase (epsps) gene and 258 bp fragment of Cry1Ab delta-endotoxin (cry1Ab) gene for GMO screening. The certified reference materials containing Roundup Ready soybean (RRS) and maize MON 810 were applied for the development and optimization of uniplex and multiplex PCR systems. Evaluation of amplification products by agarose gel electrophoresis using negative and positive controls confirmed high specificity and sensitivity at 0.1% GMO for both RRS and MON 810. The fourplex PCR was developed and optimized that allows simultaneous detection of three common transgenic elements, such as: CaMV 35S promoter, NOS terminator, epsps gene together with soybean-specific lectin gene. The triplex PCR developed enables simultaneous identification of transgenic elements, such as: 35S promoter and cry1Ab gene together with maize zein gene. The analysis of different processed foods demonstrated that multiplex PCR methods developed in this study are useful for accurate and fast screening of GM food products.

dNTP(3′‐OCH3), a 3′‐O‐methyl derivative of dNTP, is a chain terminator substrate for DNA synthesis catalyzed by AMV reverse transriptase. The enzyme seems to be the only DNA polymerase susceptible to the inhibitor while all the other DNA polymerases tested are fully resistant to the nucleotide analog. The resistant polymerases are: E. coli DNA polymerase I, Klenow's fragment of DNA polymerase I, phage T4 DNA polymerase, calf thymus DNA polymerase α, rat liver DNA polymerase β and calf thymus terminal deoxyribonucleotidyl transferase.

Background:The fis promoter upstream region harbors RNA polymerase binding sites of unknown function. Results: Modifications of the upstream polymerase binding affect fis gene expression in a supercoiling-dependent manner. Conclusion: Concomitant binding of RNA polymerase at the fis promoter and upstream region acts as a topological device regulating transcription. Significance: RNA polymerase can act as an architectural factor modulating the activity of transcription initiation complexes.