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  • 1. Baranska, S.
    et al.
    Gabig-Ciminska, Magdalena
    KTH, Superseded Departments, Biotechnology.
    Wegrzyn, A.
    Konopa, G.
    Herman-Antosiewicz, A.
    Hernandez, P.
    Schvartzman, J. B.
    Helinski, D. R.
    Wegrzyn, G.
    Regulation of the switch from early to late bacteriophage lambda DNA replication2001In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 147, p. 535-547Article in journal (Refereed)
    Abstract [en]

    There are two modes of bacteriophage lambda DNA replication following infection of its host, Escherichia coli. Early after infection, replication occurs according to the theta (theta or circle-to-circle) mode, and is later switched to the sigma (sigma or rolling-circle) mode. It is not known how this switch, occurring at a specific time in the infection cycle, is regulated. Here it is demonstrated that in type cells the replication starting from ori lambda proceeds both bidirectionally and unidirectionally, whereas in bacteria devoid of a functional DnaA protein, replication from ori lambda is predominantly unidirectional. The regulation of directionality of replication from ori lambda is mediated by positive control of lambda p(R) promoter activity by DnaA, since the mode of replication of an artificial lambda replicon bearing the p(tet) promoter instead of p(R) was found to be independent of DnaA function. These findings and results of density-shift experiments suggest that in dnaA mutants infected with lambda, phage DNA replication proceeds predominantly according to the unidirectional theta mechanism and is switched early after infection to the sigma mode. It is proposed that in wild-type E. coli cells infected with lambda, phage DNA replication proceeds according to a bidirectional theta mechanism early after infection due to efficient transcriptional activation of ori lambda, stimulated by the host DnaA protein. After a few rounds of this type of replication, the resulting increased copy number of lambda genomic DNA may cause a depletion of free DnaA protein because of its interaction with the multiple DnaA-binding sites in lambda DNA. It is proposed that this may lead to inefficient transcriptional activation of ori lambda resulting in unidirectional theta replication followed by sigma type replication.

  • 2. Barken, K. B.
    et al.
    Gabig-Ciminska, Magdalena
    KTH, Superseded Departments, Biotechnology.
    Holmgren, Anders
    KTH, Superseded Departments, Biotechnology.
    Molin, S.
    Effect of unlabeled helper probes on detection of an RNA target by bead-based sandwich hybridization2004In: BioTechniques, ISSN 0736-6205, E-ISSN 1940-9818, Vol. 36, no 1, p. 124-+Article in journal (Refereed)
    Abstract [en]

    Unlabeled helper oligonucleotides assisting a bead-based sandwich hybridization assay were tested for the optimal placement of the capture and detection probes. The target used was a full-length in vitro synthesized mRNA molecule. Helper probes complementary to regions adjacent to the binding site of the 5' end attached capture probe were found much more effective than helper probes targeting positions adjacent to the detection probe binding site. The difference is believed to be caused by a disruption of the RNA secondary structure in the area where the capture probe binds, thereby reducing structural interference from the bead. The use of additional helpers showed an additive effect. Using helpers, at both sides of the capture and detection probes showed a 15- to 40-fold increase in hybridization efficiency depending on the target, thereby increasing the sensitivity of the hybridization assays. Using an electrical chip linked to the detection probe for the detection of p-ominophenol, which is produced by alkaline phosphatase, a detection limit of 2 x 10(-13) M mRNA molecules was reached without the use of a nucleic acid amplification step.

  • 3.
    Basselet, Pascal
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Wegrzyn, Grzegorz
    Enfors, Sven-Olof
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Gabig-Ciminska, Magdalena
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Sample processing for DNA chip array-based analysis of enterohemorrhagic Escherichia coli (EHEC)2008In: Microbial Cell Factories, ISSN 1475-2859, E-ISSN 1475-2859, Vol. 7Article in journal (Refereed)
    Abstract [en]

    Background: Exploitation of DNA-based analyses of microbial pathogens, and especially simultaneous typing of several virulence-related genes in bacteria is becoming an important objective of public health these days. Results: A procedure for sample processing for a confirmative analysis of enterohemorrhagic Escherichia coli (EHEC) on a single colony with DNA chip array was developed and is reported here. The protocol includes application of fragmented genomic DNA from ultrasonicated colonies. The sample processing comprises first 2.5 min of ultrasonic treatment, DNA extraction (2x), and afterwards additional 5 min ultrasonication. Thus, the total sample preparation time for a confirmative analysis of EHEC is nearly 10 min. Additionally, bioinformatic revisions were performed in order to design PCR primers and array probes specific to most conservative regions of the EHEC-associated genes. Six strains with distinct pathogenic properties were selected for this study. At last, the EHEC chip array for a parallel and simultaneous detection of genes etpC-stx1-stx2-eae was designed and examined. This should permit to sense all currently accessible variants of the selected sequences in EHEC types and subtypes. Conclusion: In order to implement the DNA chip array-based analysis for direct EHEC detection the sample processing was established in course of this work. However, this sample preparation mode may also be applied to other types of EHEC DNA-based sensing systems.

  • 4.
    Gabig-Ciminska, Magdalena
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Developing nucleic acid-based electrical detection systems2006In: Microbial Cell Factories, ISSN 1475-2859, E-ISSN 1475-2859, Vol. 5Article in journal (Refereed)
    Abstract [en]

    Development of nucleic acid-based detection systems is the main focus of many research groups and high technology companies. The enormous work done in this field is particularly due to the broad versatility and variety of these sensing devices. From optical to electrical systems, from label-dependent to label-free approaches, from single to multi-analyte and array formats, this wide range of possibilities makes the research field very diversified and competitive. New challenges and requirements for an ideal detector suitable for nucleic acid analysis include high sensitivity and high specificity protocol that can be completed in a relatively short time offering at the same time low detection limit. Moreover, systems that can be miniaturized and automated present a significant advantage over conventional technology, especially if detection is needed in the field. Electrical system technology for nucleic acid-based detection is an enabling mode for making miniaturized to micro- and nanometer scale bio-monitoring devices via the fusion of modern micro- and nanofabrication technology and molecular biotechnology. The electrical biosensors that rely on the conversion of the Watson-Crick base-pair recognition event into a useful electrical signal are advancing rapidly, and recently are receiving much attention as a valuable tool for microbial pathogen detection. Pathogens may pose a serious threat to humans, animal and plants, thus their detection and analysis is a significant element of public health. Although different conventional methods for detection of pathogenic microorganisms and their toxins exist and are currently being applied, improvements of molecular-based detection methodologies have changed these traditional detection techniques and introduced a new era of rapid, miniaturized and automated electrical chip detection technologies into pathogen identification sector. In this review some developments and current directions in nucleic acid-based electrical detection are discussed.

  • 5.
    Gabig-Ciminska, Magdalena
    et al.
    KTH, Superseded Departments, Biotechnology.
    Andresen, Heiko
    KTH, Superseded Departments, Biotechnology.
    Albers, Joerg
    Hintsche, Rainer
    Enfors, Sven-Olof
    KTH, Superseded Departments, Biotechnology.
    Identification of pathogenic microbial cells and spores by electrochemical detection on a biochip2004In: Microbial Cell Factories, ISSN 1475-2859, E-ISSN 1475-2859, Vol. 3, p. 2-Article in journal (Refereed)
    Abstract [en]

    Background: Bacillus cereus constitutes a significant cause of acute food poisoning in humans. Despite the recent development of different detection methods, new effective control measures and better diagnostic tools are required for quick and reliable detection of pathogenic microorganisms. Thus, the objective of this study was to determine a simple method for rapid identification of enterotoxic Bacillus strains. Here, a special attention is given to an electrochemical biosensor since it meets the requirements of minimal size, lower costs and decreased power consumption. Results: A bead-based sandwich hybridization system was employed in conjugation with electric chips for detection of vegetative cells and spores of Bacillus strains based on their toxin-encoding genes. The system consists of a silicon chip based potentiometric cell, and utilizes paramagnetic beads as solid carriers of the DNA probes. The specific signals from 20 amol of bacterial cell or spore DNA were achieved in less than 4 h. The method was also successful when applied directly to unpurified spore and cell extract samples. The assay for the haemolytic enterotoxin genes resulted in reproducible signals from B. cereus and B. thuringiensis while haemolysin-negative B. subtilis strain did not yield any signal. Conclusions: The sensitivity, convenience and specificity of the system have shown its potential. In this respect an electrochemical detection on a chip enabling a fast characterization and monitoring of pathogens in food is of interest. This system can offer a contribution in the rapid identification of bacteria based on the presence of specific genes without preceding nucleic acid amplification.

  • 6.
    Gabig-Ciminska, Magdalena
    et al.
    KTH, Superseded Departments, Biotechnology.
    Herman-Antosiewicz, A.
    Kwiatkowska, M.
    Los, M.
    Thomas, M. S.
    Wegrzyn, G.
    The cell surface protein Ag43 facilitates phage infection of Escherichia coli in the presence of bile salts and carbohydrates2002In: Microbiology, ISSN 1350-0872, E-ISSN 1465-2080, Vol. 148, p. 1533-1542Article in journal (Refereed)
    Abstract [en]

    it was found that infection of Escherichia coli by bacteriophage lambda is inhibited in the presence of certain bile salts and carbohydrates when cells are in the 'OFF' state for production of the phase-variable cell surface protein antigen 43 (Ag43). The inhibition of phage growth was found to be due to a significant impairment in the process of phage adsorption. Expression of the gene encoding Ag43 (agn43) from a plasmid or inactivation of the oxyR gene (encoding an activator of genes important for defence against oxidative stress) suppressed this inhibition. A mutation, rpoA341, in the gene encoding the alpha subunit of RNA polymerase also facilitated phage adsorption in the presence of bile salts and carbohydrates. The rpoA341 mutation promoted efficient production of Ag43 in a genetic background that would otherwise be in the 'OFF' phase for expression of the agn43 gene. Analysis of a reporter gene fusion demonstrated that the promoter for the agn43 gene was more active in the rpoA341 mutant than in the otherwise isogenic rpoA(+) strain. The combined inhibitory action of bile salts and carbohydrates on phage adsorption and the abolition of this inhibition by production of Ag43 was not restricted to lambda as a similar phenomenon was observed for the coliphages P1 and T4.

  • 7.
    Gabig-Ciminska, Magdalena
    et al.
    KTH, Superseded Departments, Biotechnology.
    Holmgren, Anders
    KTH, Superseded Departments, Biotechnology.
    Andresen, Heiko
    KTH, Superseded Departments, Biotechnology.
    Barken, K. B.
    Wumpelmann, M.
    Albers, J.
    Hintsche, R.
    Breitenstein, A.
    Neubauer, P.
    Los, M.
    Czyz, A.
    Wegrzyn, G.
    Silfversparre, G.
    Jurgen, B.
    Schweder, T.
    Enfors, Sven-Olof
    KTH, Superseded Departments, Biotechnology.
    Electric chips for rapid detection and quantification of nucleic acids2004In: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 19, no 6, p. 537-546Article in journal (Refereed)
    Abstract [en]

    A silicon chip-based electric detector coupled to bead-based sandwich hybridization (BBSH) is presented as an approach to perform rapid analysis of specific nucleic acids. A microfluidic platform incorporating paramagnetic beads with immobilized capture probes is used for the biorecognition steps. The protocol involves simultaneous sandwich hybridization of a single-stranded nucleic acid target with the capture probe on the beads and with a detection probe in the reaction solution, followed by enzyme labeling of the detection probe, enzymatic reaction, and finally, potentiometric measurement of the enzyme product at the chip surface. Anti-DIG-alkaline phosphatase conjugate was used for the enzyme labeling of the DIG-labeled detection probe. p-Aminophenol phosphate (pAPP) was used as a substrate. The enzyme reaction product, p-aminophenol (pAP), is oxidized at the anode of the chip to quinoneimine that is reduced back to pAP at the cathode. The cycling oxidation and reduction of these compounds result in a current producing a characteristic signal that can be related to the concentration of the analyte. The performance of the different steps in the assay was characterized using in vitro synthesized RNA oligonucleotides and then the instrument was used for analysis of 16S rRNA in Escherichia coli extract. The assay time depends on the sensitivity required. Artificial RNA target and 16S rRNA, in amounts ranging from 10(11) to 10(10) molecules, were assayed within 25 min and 4 h, respectively.

  • 8.
    Gabig-Ciminska, Magdalena
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Liu, Yanling
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Enfors, Sven-Olof
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Gene-based identification of bacterial colonies with an electric chip2005In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 345, no 2, p. 270-276Article in journal (Refereed)
    Abstract [en]

    A method for the identification of bacterial colonies based on their content of specific genes is presented. This method does not depend on DNA separation or DNA amplification. Bacillus cereus carrying one of the genes (hblC) coding for the enterotoxin hemolysin was identified with this method. It is based on target DNA hybridization to a capturing probe immobilized on magnetic beads, followed by enzymatic labeling and measurement of the enzyme product with a silicon-based chip. An hblC-positive colony containing 10(7) cells could be assayed in 30 min after ultrasonication and centrifugation. The importance of optimizing the ultrasonication is illustrated by analysis of cell disruption kinetics and DNA fragmentation. An early endpoint PCR analysis was used to characterize the DNA fragmentation as a function of ultrasonication time. The first minutes of sonication increased the signal due to both increased DNA release and increased DNA fragmentation. The latter is assumed to increase the signal due to improved diffusion and faster hybridization of the target DNA. Too long sonication decreased the signal, presumably due to loss of hybridization sites on the targets caused by extensive DNA fragmentation. The results form a basis for rational design of an ultrasound cell disruption system integrated with analysis on chip that will move nucleic acid-based detection through real-time analysis closer to reality.

  • 9.
    Gabig-Ciminska, Magdalena
    et al.
    KTH, Superseded Departments, Biotechnology.
    Los, M.
    Holmgren, Anders
    KTH, Superseded Departments, Fibre and Polymer Technology.
    Albers, J.
    Czyz, A.
    Hintsche, R.
    Wegrzyn, G.
    Enfors, Sven-Olof
    KTH, Superseded Departments, Biotechnology.
    Detection of bacteriophage infection and prophage induction in bacterial cultures by means of electric DNA chips2004In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 324, no 1, p. 84-91Article in journal (Refereed)
    Abstract [en]

    Infections of bacterial cultures by bacteriophages are common and serious problems in many biotechnological laboratories and factories. A method for specific, quantitative, and quick detection of phage contamination, based on the use of electric DNA chip is described here. Different phages of Escherichia coli and Bacillus subtilis were analyzed. Phage DNA was isolated from bacterial culture samples and detected by combination of bead-based sandwich hybridization with enzyme-labeled probes and detection of the enzymatic product using silicon chips. The assay resulted in specific signals from all four tested phages without significant background. Although high sensitivity was achieved in 4h assay time, a useful level of sensitivity (10(7)-10(8) phages) is achievable within 25 min. A multiplex DNA chip technique involving a mixture of probes allows for detection of various types of phages in one sample.. These analyses confirmed the specificity of the assay.

  • 10.
    Gabig-Ciminska, Magdalena
    et al.
    KTH, Superseded Departments, Biotechnology.
    Wegrzyn, G.
    An introduction to DNA chips: principles, technology, applications and analysis2001In: Acta Biochimica Polonica, ISSN 0001-527X, E-ISSN 1734-154X, Vol. 48, no 3, p. 615-622Article, review/survey (Refereed)
    Abstract [en]

    This review describes the recently developed GeneChip technology that provides efficient access to genetic information using miniaturised, high-density arrays of DNA or oligonucleotide probes. Such microarrays are powerful tools to study the molecular basis of interactions on a scale that would lie impossible using conventional analysis. The recent development of the microarray technology has greatly accelerated the investigation of gene regulation. Arrays are mostly used to identify which genes are turned on or off in a cell or tissue, and also, to evaluate the extent of a gene's expression under various conditions. Indeed, this technology has been successfully applied to investigate simultaneous expression of many thousands of genes and to the detection of mutations or polymorphisms, as well as for their mapping and sequencing.

  • 11. Jasiecki, J.
    et al.
    Czyz, A.
    Gabig-Ciminska, Magdalena
    KTH, Superseded Departments, Biotechnology.
    Wegrzyn, G.
    Construction and use of a broad-host-range plasmid expressing the lamB gene for utilization of bacteriophage lambda vectors in the marine bacterium Vibrio harveyi2001In: Marine Biotechnology, ISSN 1436-2228, E-ISSN 1436-2236, Vol. 3, no 4, p. 336-345Article in journal (Refereed)
    Abstract [en]

    The remarkable success of Escherichia coli as a model organism in molecular genetics was dependent, among other things, on its susceptibility to genetic manipulation. Many versatile and sophisticated genetic tools for molecular biology studies are derived from bacteriophage lambda. However, this bacteriophage is specific for E. coli, and thus lambda -based techniques have been restricted to this bacterium. Plasmids expressing the E. coli gene coding for bacteriophage lambda receptor were reported previously, and introduction of such plasmids into cells of some other bacteria made them sensitive to phage lambda infection. However, we found that these systems were not efficient for Vibrio harveyi, one of the most frequently investigated species of marine bacteria. Here we describe construction of a broad-host-range plasmid expressing the lamB gene. Introduction of this plasmid to V. harveyi cells and expression of lamB made this strain susceptible to bacteriophage lambda adsorption and lambda DNA injection. Foreign genetic material could be introduced into cells of this strain using a cosmid vector.

  • 12.
    Liu, Yanling
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Basselet, Pascal
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Los, Marcin
    Wegrzyn, Grzegorz
    Gabig-Ciminska, Magdalena
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Enfors, Sven-Olof
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Rapid determination of virulence factors in EHEC colonies with an electric DNA chip arrayArticle in journal (Other academic)
  • 13.
    Liu, Yanling
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Elsholz, Bruno
    Fraunhofer Institute for Silicon Technology, Itzehoe, Germany.
    Enfors, Sven-Olof
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Gabig-Ciminska, Magdalena
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Confirmative electric DNA array-based test for food poisoning Bacillus cereus2007In: Journal of Microbiological Methods, ISSN 0167-7012, E-ISSN 1872-8359, Vol. 70, no 1, p. 55-64Article in journal (Refereed)
    Abstract [en]

    Detection of the full set of toxin encoding genes involved in gastrointestinal diseases caused by B. cereus was performed. Eight genes determining the B. cereus pathogenicity, which results in diarrhea or emesis, were simultaneously evaluated on a 16-position electrical chip microarray. The DNA analyte preparation procedure comprising first 5 min of ultrasonic treatment, DNA extraction, and afterwards an additional 10 min sonication, was established as the most effective way of sample processing. No DNA amplification step prior to the analysis was included. The programmed assay was carried out within 30 min, once the DNA analyte from 10(8) bacterial cells, corresponding to one agar colony, was subjected to the assay. In general, this work represents a mature analytical way for DNA review. It can be used under conditions that require almost immediate results.

  • 14.
    Liu, Yanling
    et al.
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Elsholz, Bruno
    Fraunhofer Institute for Silicon Technology, Itzehoe, Germany.
    Enfors, Sven-Olof
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Gabig-Ciminska, Magdalena
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Critical factors for the performance of chip array-based electrical detection of DNA for analysis of pathogenic bacteria2008In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 382, no 2, p. 77-86Article in journal (Refereed)
    Abstract [en]

    Different factors influencing chip array-based electrical detection of DNA for analysis of pathogenic bacteria were examined. Both rehydration of capture probe layer of functionalized chip arrays and efficient hybridization of targets irrespective of their length resulted in signal enhancement when high-ionic phosphate-buffered saline (i.e., 600 mM sodium chloride and 40 mM disodium hydrogen phosphate) was used. Similarly, placement of two adjacent capture and detection probe-binding sites at a terminal part of the target strand resulted in significant signal increase. Moreover, 10-min ultrasonic fragmentation of targets amplified the signals Lip to twofold for longer DNA strands (i.e., >300 bp). No obvious effects on signals were visible for shorter than 400-bp PCR amplicons subjected to ultrasonication. For DNA strands of all sizes, more than 10 min ultrasonication diminished the specific electrical responses. Our results also demonstrate that target analytes are detected with discrimination against mismatches even for single nucleotide sequence alteration. The mismatch detection appeared in order of ease Of recognition as follows: triple random > quintuple middle > triple middle > single middle mismatch. Among the three variants of one-base mismatches, a sequence variation was most remarkable for adenine. On the other hand, no benefits in assay sensitivity were recognized by the use of longer capture probe linkers as the 6-C linker.

  • 15.
    Wang, Yong-Bin
    et al.
    KTH, School of Information and Communication Technology (ICT), Integrated Devices and Circuits.
    Assefaw-Redda, Yohannes
    KTH, School of Biotechnology (BIO).
    Gabig-Ciminska, Magdalena
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Enfors, Sven-Olof
    KTH, School of Biotechnology (BIO), Bioprocess Technology.
    Östling, Mikael
    KTH, School of Information and Communication Technology (ICT), Microelectronics and Information Technology, IMIT.
    Zhang, Shi-Li
    KTH, School of Information and Communication Technology (ICT), Microelectronics and Information Technology, IMIT.
    A novel dual mode capacitor biosensor for real-time, label-free DNA detection2006In: 2006 INTERNATIONAL ELECTRON DEVICES MEETING, VOLS 1 AND 2, NEW YORK: IEEE , 2006, p. 447-450Conference paper (Refereed)
    Abstract [en]

    A novel biosensor working in both MOS capacitor and electric double-layer capacitor mode for real-time, label-free DNA detection is presented. The former mode measures density and polarity of charged biomolecules to be monitored, while the latter mode appears to reveal more of surface-molecule interactions and offers high sensitivity and easiness for diagnosis. A combined mode-utilization yields optimal complementary information of significance.

  • 16. Wegrzyn, A.
    et al.
    Czyz, A.
    Gabig-Ciminska, Magdalena
    KTH, Superseded Departments, Biotechnology.
    Wegrzyn, G.
    ClpP/ClpX-mediated degradation of the bacteriophage lambda O protein and regulation of lambda phage and lambda plasmid replication2000In: Archives of Microbiology, ISSN 0302-8933, E-ISSN 1432-072X, Vol. 174, no 02-jan, p. 89-96Article in journal (Refereed)
    Abstract [en]

    The O protein is a replication initiator that binds to the ori lambda region and promotes assembly of the bacteriophage lambda replication complex. This protein, although protected from proteases by other elements of the replication complex, in a free form is rapidly degraded in the host, Escherichia coil, by the ClpP/ClpX protease. Nevertheless, the physiological role of this rapid degradation remains unclear. Here we demonstrate that the copy number of plasmids derived from bacteriophage lambda is significantly higher in wild-type cells growing in rich media than in slowly growing bacteria. However, lambda plasmid copy number in bacteria devoid of the ClpP/ClpX protease was not dependent on the bacterial growth rate and in all minimal media rested was comparable to that observed in wildtype cells growing in a rich medium. Contrary to lambda plasmid replication, the efficiency of lytic growth of bacteriophage lambda was found to be dependent on the host growth rate in both wild-type bacteria and clpP and clpX mutants. The activities of two major lambda promoters operating during the lytic development, p(R) and p(L), were found to be slightly dependent on the host growth rate. However, when p(R) activity was significantly decreased in the dnaA mutant, production of phage progeny was completely abolished at low growth rates. These results indicate that the O protein (whose level in E. coli cells depends on the activity of ClpP/ClpX protease) is a major limiting factor in the regulation of lambda plasmid replication at low bacterial growth rates. However, this protein seems to be only one of the limiting factors in the bacteriophage lambda lytic development under poor growth conditions of host cells. Therefore, it seems that the role of the rapid ClpP/ClpX-mediated proteolysis of the O protein is to decrease the efficiency of early DNA replication of the phage in slowly growing host cells.

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