Publications

analysis of recombinant protein toxicity in e. coli through a phage lambda-based genetic screening system.	the aspartic protease from the human immunodeficiency virus type 1 (hiv-1) is highly toxic to e. coli, thus impairing its yield in production processes. proteolytic cleavage of essential cellular proteins is probably a major contributor to the bacteriocidal effect but this has not been proven. through an adapted high-throughput lambda-based screening system, we have analyzed a set of hiv-1 protease mutants with distinguishable catalytic properties and we show that inactive enzymes are as toxic t ...	2007	17479218
incorporation of a lambda phage recombination system and egfp detection to simplify mutagenesis of herpes simplex virus bacterial artificial chromosomes.	targeted mutagenesis of the herpesvirus genomes has been facilitated by the use of bacterial artificial chromosome (bac) technology. such modified genomes have potential uses in understanding viral pathogenesis, gene identification and characterization, and the development of new viral vectors and vaccines. we have previously described the construction of a herpes simplex virus 2 (hsv-2) bac and the use of an allele replacement strategy to construct hsv-2 recombinants. while the bac mutagenesis ...	2007	17501993
strand opening-deficient escherichia coli rna polymerase facilitates investigation of closed complexes with promoter dna: effects of dna sequence and temperature.	formation of the strand-separated, open complex between rna polymerase and a promoter involves several intermediates, the first being the closed complex in which the dna is fully base-paired. this normally short lived complex has been difficult to study. we have used a mutant escherichia coli rna polymerase, deficient in promoter dna melting, and variants of the p(r) promoter of bacteriophage lambda to model the closed complex intermediate at physiologically relevant temperatures. our results in ...	2007	17507375
general transfer matrix formalism to calculate dna-protein-drug binding in gene regulation: application to or operator of phage lambda.	the transfer matrix methodology is proposed as a systematic tool for the statistical-mechanical description of dna-protein-drug binding involved in gene regulation. we show that a genetic system of several cis-regulatory modules is calculable using this method, considering explicitly the site-overlapping, competitive, cooperative binding of regulatory proteins, their multilayer assembly and dna looping. in the methodological section, the matrix models are solved for the basic types of short- and ...	2007	17526526
plasmids derived from gifsy-1/gifsy-2, lambdoid prophages contributing to the virulence of salmonella enterica serovar typhimurium: implications for the evolution of replication initiation proteins of lambdoid phages and enterobacteria.	gifsy-1 and gifsy-2 are lambdoid prophages which contribute to the virulence of salmonella enterica serovar typhimurium. the nucleotide sequence of the replication region of both prophages is identical, and similar in organization to the replication region of bacteriophage lambda. to investigate the replication of the gifsy phages and the relationship between gifsy and host chromosome replication, a plasmid which contained all the genes and regulatory sequences required for autonomous replicatio ...	2007	17526845
trans cooperativity by a split dna recombinase: the central and catalytic domains of bacteriophage lambda integrase cooperate in cleaving dna substrates when the two domains are not covalently linked.	site-specific recombinases of the lambda-integrase family recognize and cleave their cognate dna sites through cooperative binding to opposite sides of the dna substrate by a c-terminal catalytic domain and a flexibly linked "core-binding" domain; regulation of this cleavage is achieved via the formation of higher-order complexes. we report that the core-binding domain of lambda-integrase is able to stimulate the activity of the catalytic domain even when the two domains are not linked. this tra ...	2007	17531268
internal dna pressure modifies stability of wt phage.	dsdna in bacteriophages is highly stressed and exerts internal pressures of many atmospheres (1 atm = 101.3 kpa) on the capsid walls. we investigate the correlation between packaged dna length in lambda phage (78-100% of wt dna) and capsid strength by using an atomic force microscope indentation technique. we show that phages with wt dna are twice as strong as shorter genome mutants, which behave like empty capsids, regardless of high internal pressure. our analytical model of dna-filled capsid ...	2007	17535894
the crystal structure of lambda-gam protein suggests a model for recbcd inhibition.	in escherichia coli, recbcd processes double-stranded dna breaks during the initial stages of homologous recombination. recbcd contains helicase and nuclease activities, and unwinds and digests the blunt-ended dna until a specific eight-nucleotide sequence, chi, is encountered. chi modulates the nuclease activity of recbcd and results in a resected dna end, which is a substrate for reca during subsequent steps in recombination. recbcd also acts as a defence mechanism against bacteriophage infect ...	2007	17544443
crystallization and preliminary crystallographic characterization of the origin-binding domain of the bacteriophage lambda o replication initiator.	the bacteriophage lambda o protein binds to the lambda replication origin (orilambda) and serves as the primary replication initiator for the viral genome. the binding energy derived from the binding of o to orilambda is thought to help drive dna opening to facilitate initiation of dna replication. detailed understanding of this process is severely limited by the lack of high-resolution structures of o protein or of any lambdoid phage-encoded paralogs either with or without dna. the production o ...	2007	17554183
ionic effects on viral dna packaging and portal motor function in bacteriophage phi 29.	in many viruses, dna is confined at such high density that its bending rigidity and electrostatic self-repulsion present a strong energy barrier in viral assembly. therefore, a powerful molecular motor is needed to package the dna into the viral capsid. here, we investigate the role of electrostatic repulsion on single dna packaging dynamics in bacteriophage phi 29 via optical tweezers measurements. we show that ionic screening strongly affects the packing forces, confirming the importance of el ...	2007	17556543
mapping the tip-sample interactions on dppc and dna by dynamic force spectroscopy under ambient conditions.	self-assembled monolayers of dppc and dna adsorbed on mica are examined by dynamic force spectroscopy under ambient conditions. by a systematic recording of the frequency shift caused by the tip-sample interaction we determine the corresponding tip-sample potential and force curves. in difference to the conventional measurement of force-vs.-distance curves this technique allows the continuous measurement of tip-sample forces without instabilities caused by a jump-to-contact. due to the systemati ...	2007	17566660
modifications enhance the apoptosis-inducing activity of fadd.	the ability to enhance apoptosis-inducing activity in specific cells, despite the presence of cellular antiapoptotic proteins, would allow the removal of target cells from a cell population. here, we show that modification of fas-associated protein with death domain (fadd) by fusing the tandem death effector domains (ded) of fadd to the e protein of lambda phage, a head coat protein with self-assembly activity, greatly increases the apoptosis-inducing activity of fadd in both adherent nih3t3 and ...	2007	17575108
[construction of a cdna library from liver tissue of rhesus monkey, macaca mulatta].	to screen the target rhesus genes and give some basic genetic evidences to its value as one of the most important animal model in biomedical study, we constructed a cdna expression library from liver tissue of a healthy rhesus monkey.	2007	17593811
interaction of the gifsy-1 xis protein with the gifsy-1 attp sequence.	the gifsy-1 phage integrates site specifically into the salmonella chromosome via an integrase-mediated site-specific recombination mechanism. initial genetic analysis suggests that gifsy-1 integrase-mediated excision of the gifsy-1 phage is influenced by proteins encoded by both the gifsy-1 and the gifsy-2 phages. our studies show that the gifsy-1 xis protein regulates the directionality of integrase-mediated excision of the gifsy-1 phage. electrophoretic mobility shift assays, dnase i footprin ...	2007	17601790
lambdan-gfp: an rna reporter system for live-cell imaging.	we describe a gfp-based rna reporter system (lambdan-gfp) to visualize rna molecules in live mammalian cells. it consists of gfp fused to an arginine-rich peptide derived from the phage lambda n protein, lambdan22, which binds a unique minimal rna motif and can be used to tag any rna molecule. lambdan-gfp uses a small and easy to engineer rna tag, reducing the likelihood of perturbing the function of the tagged rna molecule.	2007	17603490
modelling the stability of stx lysogens.	shiga-toxin-converting bacteriophages (stx phages) are temperate phages of escherichia coli, and can cause severe human disease. the spread of shiga toxins by stx phages is directly linked to lysogen stability because toxins are only synthesized and released once the lytic cycle is initiated. lysogens of stx phages are known to be less stable than those of the related lambda phage; this is often described in terms of a 'hair-trigger' molecular switch from lysogeny to lysis. we have developed a m ...	2007	17604057
characterization of the dsdna prophage sequences in the genome of neisseria gonorrhoeae and visualization of productive bacteriophage.	bioinformatic analysis of the genome sequence of neisseria gonorrhoeae revealed the presence of nine probable prophage islands. the distribution, conservation and function of many of these sequences, and their ability to produce bacteriophage particles are unknown.	2007	17615066
symbolic modeling of genetic regulatory networks.	understanding the functioning of genetic regulatory networks supposes a modeling of biological processes in order to simulate behaviors and to reason on the model. unfortunately, the modeling task is confronted to incomplete knowledge about the system. to deal with this problem we propose a methodology that uses the qualitative approach developed by thomas. a symbolic transition system can represent the set of all possible models in a concise and symbolic way. we introduce a new method based on ...	2007	17636866
noncanonical interactions in the management of rna structural blocks by the transcription termination rho helicase.	to trigger transcription termination, the ring-shaped rna-dna helicase rho from escherichia coli chases the rna polymerase along the nascent transcript, starting from a single-stranded c-rich rut (rho utilization) loading site. in some instances, a small hairpin structure divides harmlessly the c-rich loading region into two smaller rut subsites, best exemplified by the tr1 terminator from phage lambda. here, we show that the rho helicase can also elude a rna structural block located far downstr ...	2007	17655325
measurements of the hysteresis in unzipping and rezipping double-stranded dna.	complete unzipping and rezipping of lambda -phage double-stranded dna is achieved by applying a constant force. a strong hysteresis is observed at all tested time scales and temperatures. hysteresis also occurs for partial unzipping, indicating stability for the partially open state over a force range of 2- 5pn . results are compared to nearest-neighbor model simulations, and reasonable agreement is found.	2007	17677099
dynamic and static light scattering analysis of dna ejection from the phage lambda.	with the aid of time-resolved dynamic light scattering (dls) and static light scattering (sls), we have analyzed the ejection kinetics from the bacterial virus bacteriophage (or phage) lambda , triggered in vitro by its receptor. we have used dls to investigate the kinetics in such a system. furthermore, we have shown that both sls and dls can be interchangeably used to study the process of phage dna release. dls is superior to sls in that it also allows the change in the light scattering arisin ...	2007	17677501
a study of comparability in amplified fragment length polymorphism profiling using a simple model system.	a simple amplified fragment length polymorphism (aflp) model, using the bacteriophage lambda genome, was developed to test the reproducibility of this technique in an international comparative study. using either non-selective or selective primers, nine fragments or subsets of two or three fragments, respectively, were predicted using in silico software. under optimized conditions, all predicted fragments were experimentally generated. the reproducibility of the aflp model was tested by submitti ...	2007	17696213
global analysis of host response to induction of a latent bacteriophage.	the transition from viral latency to lytic growth involves complex interactions among host and viral factors, and the extent to which host physiology is buffered from the virus during induction of lysis is not known. a reasonable hypothesis is that the virus should be evolutionarily selected to ensure host health throughout induction to minimize its chance of reproductive failure. to address this question, we collected transcriptional profiles of escherichia coli and bacteriophage lambda through ...	2007	17764558
genome sequence comparison and superinfection between two related pseudomonas aeruginosa phages, d3112 and mp22.	a temperate transposable bacteriophage (mp22) was isolated from a korean clinical isolate of pseudomonas aeruginosa. it has a coliphage lambda-like morphology and a double-stranded dna genome. the complete nucleotide sequence and annotation of the mp22 genome and its characteristics are presented. the mp22 genome is 36 409 bp long with a g+c content of 64.2 mol%. the genome contains 51 proposed orfs, of which 48 (94 %) display synteny and significant nucleotide and protein sequence similarity to ...	2007	17768233
real-time observations of single bacteriophage lambda dna ejections in vitro.	the physical, chemical, and structural features of bacteriophage genome release have been the subject of much recent attention. many theoretical and experimental studies have centered on the internal forces driving the ejection process. recently, mangenot et al. [mangenot s, hochrein m, rädler j, letellier l (2005) curr biol 15:430-435.] reported fluorescence microscopy of phage t5 ejections, which proceeded stepwise between dna nicks, reaching a translocation speed of 75 kbp/s or higher. it is ...	2007	17804798
erratum.	in the report by k. denniston-thompson et al., entitled "physical structure of the replication origin of bacteriophage lambda" (9 december 1977, pp. 1051-1056), the t.a base pair at position 1426 (fig. 6) should be an a.t base pair; also the position of the g.c base pair affected by the til2 mutation should be 1453 rather than 1451.	1978	17840776
the dna maturation domain of gpa, the dna packaging motor protein of bacteriophage lambda, contains an atpase site associated with endonuclease activity.	terminase enzymes are common to double-stranded dna (dsdna) viruses and are responsible for packaging viral dna into the confines of an empty capsid shell. in bacteriophage lambda the catalytic terminase subunit is gpa, which is responsible for maturation of the genome end prior to packaging and subsequent translocation of the matured dna into the capsid. dna packaging requires an atpase catalytic site situated in the n terminus of the protein. a second atpase catalytic site associated with the ...	2007	17870092
probing the antiprotease activity of lambdaciii, an inhibitor of the escherichia coli metalloprotease hflb (ftsh).	the ciii protein encoded by the temperate coliphage lambda acts as an inhibitor of the ubiquitous escherichia coli metalloprotease hflb (ftsh). this inhibition results in the stabilization of transcription factor lambdacii, thereby helping the phage to lysogenize the host bacterium. lambdaciii, a small (54-residue) protein of unknown structure, also protects sigma(32), another specific substrate of hflb. in order to understand the details of the inhibitory mechanism of ciii, we cloned and expres ...	2007	17890311
effects of salt concentrations and bending energy on the extent of ejection of phage genomes.	recent work has shown that pressures inside dsdna phage capsids can be as high as many tens of atmospheres; it is this pressure that is responsible for initiation of the delivery of phage genomes to host cells. the forces driving ejection of the genome have been shown to decrease monotonically as ejection proceeds, and hence to be strongly dependent on the genome length. here we investigate the effects of ambient salts on the pressures inside phage-lambda, for the cases of mono-, di-, and tetrav ...	2008	17890396
mad-ct-2 identified as a novel melanoma cancer-testis antigen using phage immunoblot analysis.	one focus in the field of tumor immunology is the identification of cancer-specific antigens that might be exploited as therapeutic targets or as immunologic diagnostic markers. cancer-testis antigens (ctas) are of particular interest as potential target antigens given that their expression is typically restricted to germ cells among normal tissues, but aberrantly expressed in multiple tumor types. in the current report, we sought to evaluate serum antibody immune responses to a defined panel of ...	2007	17893560
temperature distribution effects on micro-cfpcr performance.	continuous flow polymerase chain reactors (cfpcrs) are biomems devices that offer unique capabilities for the ultra-fast amplification of target dna fragments using repeated thermal cycling, typically over the following temperature ranges: 90 degrees c-95 degrees c for denaturation, 50 degrees c-70 degrees c for renaturation, and 70 degrees c-75 degrees c for extension. in cfpcr, dna cocktail is pumped through the constant temperature zones and reaches thermal equilibrium with the channel walls ...	2008	17896180
chromosomal model for analysis of a long ctg/cag tract stability in wild-type escherichia coli and its nucleotide excision repair mutants.	many human hereditary neurological diseases, including fragile x syndrome, myotonic dystrophy, and friedreich's ataxia, are associated with expansions of the triplet repeat sequences (trs) (cgg/ccg, ctg/cag, and gaa/ttc) within or near specific genes. mechanisms that mediate mutations of trs include dna replication, repair, and gene conversion and (or) recombination. the involvement of the repair systems in trs instability was investigated in escherichia coli on plasmid models, and the results s ...	2007	17898841
identification of dna-binding protein target sequences by physical effective energy functions: free energy analysis of lambda repressor-dna complexes.	specific binding of proteins to dna is one of the most common ways gene expression is controlled. although general rules for the dna-protein recognition can be derived, the ambiguous and complex nature of this mechanism precludes a simple recognition code, therefore the prediction of dna target sequences is not straightforward. dna-protein interactions can be studied using computational methods which can complement the current experimental methods and offer some advantages. in the present work w ...	2007	17900341
rz/rz1 lysis gene equivalents in phages of gram-negative hosts.	under usual laboratory conditions, lysis by bacteriophage lambda requires only the holin and endolysin genes, but not the rz and rz1 genes, of the lysis cassette. defects in rz or rz1 block lysis only in the presence of high concentrations of divalent cations. the lambda rz and rz1 lysis genes are remarkable in that rz1, encoding an outer membrane lipoprotein, is completely embedded in the +1 register within rz, which itself encodes an integral inner membrane protein. while rz and rz1 equivalent ...	2007	17900620
development of a bacterial cloning vector for expression of scorpion toxins for biotechnological studies.	scorpion venoms contain toxic peptides that recognize k(+) channels of excitable and non-excitable cells. these toxins comprise three structurally distinct groups designated alpha-ktx, beta-ktx, and gamma-ktx. it is highly desirable to develop systems for the expression of these toxins for further physiological and structural studies. in this work, an expression vector (ptev3) was constructed by inserting protein d (major capsid of phage lambda) and tev protease recognition site into plasmid pet ...	2008	17904381
cro's role in the ci cro bistable switch is critical for {lambda}'s transition from lysogeny to lytic development.	ci represses cro; cro represses ci. this double negative feedback loop is the core of the classical ci-cro epigenetic switch of bacteriophage lambda. despite the classical status of this switch, the role in lambda development of cro repression of the p(rm) promoter for ci has remained unclear. to address this, we created binding site mutations that strongly impaired cro repression of p(rm) with only minimal effects on ci regulation of p(rm). these mutations had little impact on lambda developmen ...	2007	17908932
measurements of single dna molecule packaging dynamics in bacteriophage lambda reveal high forces, high motor processivity, and capsid transformations.	molecular motors drive genome packaging into preformed procapsids in many double-stranded (ds)dna viruses. here, we present optical tweezers measurements of single dna molecule packaging in bacteriophage lambda. dna-gpa-gpnu1 complexes were assembled with recombinant gpa and gpnu1 proteins and tethered to microspheres, and procapsids were attached to separate microspheres. dna binding and initiation of packaging were observed within a few seconds of bringing these microspheres into proximity in ...	2007	17919653
distinct human prolactin (hprl) and growth hormone (hgh) behavior under bacteriophage lambda pl promoter control: temperature plays a major role in protein yields.	when producing recombinant protein for therapy, it is desirable not only to obtain substantial amounts of the protein, but also to make sure that potential contaminants such as inducing agents are not present in the final product. to prevent this, one can use expression systems in which the promoter (lambdap(l)) is activated by a temperature shift that denatures a repressor (e.g., cits). in this manner, hgh was successfully expressed and secreted in escherichia coli periplasm, with specific yiel ...	2008	17920153
glycine n-methyltransferase-/- mice develop chronic hepatitis and glycogen storage disease in the liver.	glycine n-methyltransferase (gnmt) affects genetic stability by regulating dna methylation and interacting with environmental carcinogens. to establish a gnmt knockout mouse model, 2 lambda phage clones containing a mouse gnmt genome were isolated. at 11 weeks of age, the gnmt-/- mice had hepatomegaly, hypermethioninemia, and significantly higher levels of both serum alanine aminotransferase and hepatic s-adenosylmethionine. such phenotypes mimic patients with congenital gnmt deficiencies. a rea ...	2007	17937387
regulation of excision of integrative and potentially conjugative elements from streptococcus thermophilus: role of the arp1 repressor.	the integrative and conjugative elements (ices) excise by site-specific recombination between attl and attr flanking sites, self-transfer the resulting circular form and integrate into the genome of the recipient cell. two putative ices, icest1 and icest3, are integrated in the same locus in 2 strains of streptococcusthermophilus. icest1 is a composite element harbouring an internal recombination site, attl'. the recombination between attl' and attr leads to the excision of a shorter putative ic ...	2008	17957106
cooperative dna binding by ci repressor is dispensable in a phage lambda variant.	complex gene regulatory circuits contain many interacting components. in principle, all of these components and interactions may be essential to the function of the circuit. alternatively, some of them may be refinements to a simpler version of the circuit that improve its fitness. in this work, we have tested whether a particular property of a critical regulatory protein, ci, is essential to the behavior of the phage lambda regulatory circuit. in the lysogenic state, ci represses the expression ...	2007	17962420
recombination-ready sindbis replicon expression vectors for transgene expression.	sindbis viruses have been widely used as tools to study gene function in cells. despite the utility of these systems, the construction and production of alphavirus replicons is time consuming and inefficient due to potential additional restriction sites within the insert region and lack of directionality for insert ligation. in this report, we present a system useful for producing recombinant sindbis replicons that uses lambda phage recombination technology to rapidly and specifically construct ...	2007	17963504
wheat endonuclease wen1 dependent on s-adenosyl-l-methionine and sensitive to dna methylation status.	ca(2+)-, mg(2+)-dependent wheat endonuclease wen1 with molecular mass of about 27 kda was isolated from coleoptyles. methylated dna of lambda phage grown on e. coli dam(+), dcm(+) cells was hydrolyzed by wen1 more effectively than dna of phage grown on dam(-), dcm(-) cells. two ph activity maxima (ph 6.5-7.5 and 9.0-10.5) were observed when double-stranded dna was hydrolyzed. wen1 is stable at elevated temperatures (65 degrees c ) and in wide range of ph values. wen1 is activated by s-adenosyl-l ...	2007	17965597
an allosteric intramolecular pdz-pdz interaction modulates ptp-bl pdz2 binding specificity.	pdz (acronym of the synapse-associated protein psd-95/sap90, the septate junction protein discs-large, and the tight junction protein zo-1) domains are abundant small globular protein interaction domains that mainly recognize the carboxyl termini of their target proteins. detailed knowledge on pdz domain binding specificity is a prerequisite for understanding the interaction networks they establish. we determined the binding preference of the five pdz domains in the protein tyrosine phosphatase ...	2007	17979300
removal of deoxyinosine from the escherichia coli chromosome as studied by oligonucleotide transformation.	deoxyinosine (di) is produced in dna by the hydrolytic or nitrosative deamination of deoxyadenosine. it is excised in a repair pathway that is initiated by endonuclease v, the product of the nfi gene. the repair was studied in vivo using high-efficiency oligonucleotide transformation mediated by the beta protein of bacteriophage lambda in a mismatch repair-deficient host. escherichia coli was transformed with oligonucleotides containing a selectable a-g base substitution mutation. when the mutag ...	2008	17981100
characterization of the 2',3' cyclic phosphodiesterase activities of clostridium thermocellum polynucleotide kinase-phosphatase and bacteriophage lambda phosphatase.	clostridium thermocellum polynucleotide kinase-phosphatase (cthpnkp) catalyzes 5' and 3' end-healing reactions that prepare broken rna termini for sealing by rna ligase. the central phosphatase domain of cthpnkp belongs to the dinuclear metallophosphoesterase superfamily exemplified by bacteriophage lambda phosphatase (lambda-pase). cthpnkp is a ni(2+)/mn(2+)-dependent phosphodiesterase-monoesterase, active on nucleotide and non-nucleotide substrates, that can be transformed toward narrower meta ...	2007	17986465
fluorescent reference strains of bacteria by chromosomal integration of a modified green fluorescent protein gene.	fluorescent reference strains of bacteria carrying a stable chromosomally integrated single copy of the gfp gene have been developed. a modified version of the gfp gene has been generated by mutagenesis and expressed under the control of the bacteriophage lambda promoter p(l). a cassette comprising bacteriophage mu transposon arms flanking the modified gfp gene and regulatory regions was irreversibly integrated as an in-vitro-assembled transposition complex into the genomes of escherichia coli a ...	2008	17994234
incremental and unifying modelling formalism for biological interaction networks.	an appropriate choice of the modeling formalism from the broad range of existing ones may be crucial for efficiently describing and analyzing biological systems.	2007	17996051
dna recognition via mutual-induced fit by the core-binding domain of bacteriophage lambda integrase.	bacteriophage lambda integrase (lambda-int), a phage-encoded dna recombinase, cleaves its substrate dna to facilitate the formation and later resolution of a holliday junction intermediate during recombination. the core-binding and catalytic domains of lambda-int constitute a bipartite enzyme that mediates site-specific dna cleavage through their interactions with opposite sides of the recognition sequence. despite minimal direct contact between the domains, the core-binding domain has been show ...	2007	18001133
identification and characterization of int (integrase), xis (excisionase) and chromosomal attachment sites of the integrative and conjugative element icebs1 of bacillus subtilis.	icebs1 is an integrative and conjugative element (conjugative transposon) integrated into trns-leu2 in bacillus subtilis. in response to dna damage or high concentrations of potential mating partners, icebs1 can excise and transfer to various recipients, including other species. we found that excision of icebs1 occurs by site-specific recombination within 60 bp direct repeats that mark the junctions between icebs1 and chromosomal dna. excision required two icebs1 genes, int (integrase, ydcl), pr ...	2007	18005101
noise-induced switches in network systems of the genetic toggle switch.	bistability, the capacity to achieve two distinct stable steady states in response to a set of external stimuli, arises within biological systems ranging from the lambda phage switch in bacteria to cellular signal transduction pathways in mammalian cells. on the other hand, more and more experimental evidence in the form of bimodal population distribution has indicated that noise plays a very important role in the switching of bistable systems. however, the physiological mechanism underling nois ...	2007	18005421
an integrated optics microfluidic device for detecting single dna molecules.	a fluorescence-based integrated optics microfluidic device is presented, capable of detecting single dna molecules in a high throughput and reproducible manner. the device integrates microfluidics for dna stretching with two optical elements for single molecule detection (smd): a plano-aspheric refractive lens for fluorescence excitation (illuminator) and a solid parabolic reflective mirror for fluorescence collection (collector). although miniaturized in size, both optical components were produ ...	2007	18030399
measurement of the salt-dependent stabilization of partially open dna by escherichia coli ssb protein.	the rezipping force of two complementary dna strands under tension has been measured in the presence of escherichia coli single-stranded-binding proteins under salt conditions ranging from 10- to 400 mm nacl. the effectiveness of the binding protein in preventing rezipping is strongly dependent on salt concentration and compared with the salt dependence in the absence of the protein. at concentrations less than 50 mm nacl, the protein prevents complete rezipping of lambda-phage on the 2-s timesc ...	2008	18032436
physical analyses of e. coli heteroduplex recombination products in vivo: on the prevalence of 5' and 3' patches.	homologous recombination in escherichia coli creates patches (non-crossovers) or splices (half crossovers), each of which may have associated heteroduplex dna. heteroduplex patches have recombinant dna in one strand of the duplex, with parental flanking markers. which dna strand is exchanged in heteroduplex patches reflects the molecular mechanism of recombination. several models for the mechanism of e. coli recbcd-mediated recombinational double-strand-end (dse) repair specify that only the 3'- ...	2007	18043749
identification of multiple integration sites for stx-phage phi24b in the escherichia coli genome, description of a novel integrase and evidence for a functional anti-repressor.	the key virulence factor in shiga-toxigenic escherichia coli is the expression of shiga toxin (stx), which is conferred by stx-encoding temperate lambdoid phages (stx-phages). it had been assumed that stx-phages would behave similarly to lambda phage. however, contrary to the lambda superinfection immunity model, it has been demonstrated that double lysogens can be produced with the stx-phage phi24(b). here, the phi24(b) integrase gene is identified, and the preferred site of integration defined ...	2007	18048923
development of a simple cell lysis method for recombinant dna using bacteriophage lambda lysis genes.	in this study, we describe the development of a simple and efficient method for cell lysis via the insertion of a bacteriophage lambda lysis gene cluster into the pet22b expression vector in the following order; the t7 promoter, a gene for a target protein intended for production, sam7 and r. this insertion of r and sam7 into pet22b exerted no detrimental effects on cellular growth or the production of a target protein. the induction of the t7 promoter did not in itself result in the autolysis o ...	2007	18176547
a phage display system designed to detect and study protein-protein interactions.	analysing protein-protein interactions is critical in proteomics and drug discovery. the usage of 2-hybrid (2lambda) systems is limited to an in vivo environment. we describe a bacteriophage 2-hybrid system for studying protein interactions in vitro. bait and prey are displayed as fusions to the surface of phage lambda that are marked with different selectable drug-resistant markers. an interaction of phages in vitro through displayed proteins allows bacterial infection by two phages resulting i ...	2008	18179417
fc receptor-mediated, antibody-dependent enhancement of bacteriophage lambda-mediated gene transfer in mammalian cells.	lambda phage vectors mediate gene transfer in cultured mammalian cells and in live mice, and in vivo phage-mediated gene expression is increased when mice are pre-immunized with bacteriophage lambda. we now show that, like eukaryotic viruses, bacteriophage vectors are subject to fc receptor-mediated, antibody-dependent enhancement of infection in mammalian cells. antibody-dependent enhancement of phage gene transfer required fcgammari, but not its associated gamma-chain, and was not supported by ...	2008	18191979
dna bending in transcription initiation.	electrophoretic mobility shift (bandshift) phasing analysis and rotational variant topological analysis were performed on initiation complexes formed on the bacteriophage lambda pr promoter. both the open complex and an abortive complex containing a short rna primer extending to +3 were characterized. the two methods were used to analyze a series of constructs containing tandemly repeated copies of the pr promoter, with the repeat length increased in single base pair increments to progressively ...	2008	18205392
dna bubble formation in transcription initiation.	the properties of the dna bubble in the transcription open complex have been characterized by topological analysis of dna circles containing the lac uv5 promoter or the pr promoter from bacteriophage lambda. topological analysis is particularly well suited to this purpose since it quantifies the changes in dna duplex geometry caused by bubble formation as well as by superhelical dna wrapping. the duplex unwinding that results from bubble formation is detected as a reduction in topological linkin ...	2008	18205393
efficient point mutagenesis in mycobacteria using single-stranded dna recombineering: characterization of antimycobacterial drug targets.	construction of genetically isogenic strains of mycobacteria is complicated by poor recombination rates and the lack of generalized transducing phages for mycobacterium tuberculosis. we report here a powerful method for introducing single point mutations into mycobacterial genomes using oligonucleotide-derived single-stranded dna recombineering and mycobacteriophage-encoded proteins. phage che9c gp61-mediated recombination is sufficiently efficient that single base changes can be introduced with ...	2008	18221264
the c-terminal domain of escherichia coli yfhd functions as a lytic transglycosylase.	the hypothetical escherichia coli protein yfhd has been identified as the archetype for the family 1b lytic transglycosylases despite a complete lack of experimental characterization. the yfhd gene was amplified from the genomic dna of e. coli w3110 and cloned to encode a fusion protein with a c-terminal his(6) sequence. the enzyme was found to be localized to the outer membrane of e. coli, as would be expected for a lytic transglycosylase. its gene was engineered for the production of a truncat ...	2008	18234673
use of the lambda red recombinase system to rapidly generate mutants in pseudomonas aeruginosa.	the red recombinase system of bacteriophage lambda has been used to inactivate chromosomal genes in various bacteria and fungi. the procedure consists of electroporating a polymerase chain reaction (pcr) fragment that was obtained with a 1- or 3-step pcr protocol and that carries an antibiotic cassette flanked by a region homologous to the target locus into a strain that expresses the lambda red recombination system.	2008	18248677
dna molecule dynamics in converging-diverging microchannels.	the conformation and dynamics of double-stranded dna molecules in a pressure-driven flow-through sharp/gradual converging (contraction half)-diverging (expansion half) square microchannels with a contraction/expansion ratio of 4:1/1:4 were examined using fluorescently labelled lambda-phage dna at 2.2< or =de< or =30.7 (de is deborah number, a dimensionless number used in rheology to characterize how fluid a material is). both the diffusion and stretching of individual dna molecules and their loc ...	2009	18251714
plating lambda phage to generate plaques.	this unit provides detailed protocols for isolating a single plaque by titering serial dilutions or streaking on a lawn of cells. a discussion of phage transfection and in vitro packaging is also included.	2001	18265039
expression using vectors with phage lambda regulatory sequences.	in the expression system described here, plasmids (pskf) utilize regulatory signals--such as the powerful promoter pl--from the bacteriophage lambda. transcription from pl can be fully repressed and plasmids containing it are thus stabilized by the lambda repressor, ci. the repressor is supplied by an e. coli host which contains a integrated copy of a portion of the lambda genome. this so-called defective lysogen supplies the lambda regulatory proteins ci and n but does not provide the lytic com ...	2001	18265129
genomic dna libraries.	genomic dna libraries are almost always screened by hybridization using a radioactive nucleic acid probe. since this approach is essentially independent of a particular vector or type of target dna, the main problem faced when considering creation of a genomic dna library is simply generating a large enough number of recombinant dna clones. the basic strategies used to address this problem have included both minimizing the number of clones necessary by incorporating large fragments of genomic dn ...	2001	18265243
recombination-based assay (rba) for screening bacteriophage lambda libraries.	the recombination-based assay represents a convenient way to screen a complex library constructed in bacteriophage lambda for homology to a given sequence cloned into a specially designed plasmid. the technique serves to screen a bacteriophage library rapidly and efficiently with a sequence cloned into a plasmid; counterselection then yields the gene product of interest with its plasmid carrier deleted. because 10(6) to 10(7) plaque-forming units (pfu) may be screened using several petri dishes, ...	2001	18265256
preparation of templates for dna sequencing.	this unit contains protocols for preparing dna suitable for use as dideoxy sequencing templates and as material for end labeling and chemical sequencing. in all protocols, the starting material contains the recombinant molecule to be sequenced. dna from m13mp-derived phage is easily prepared and is currently the most reliable source of template for large-scale dideoxy sequencing projects. because it is occasionally necessary or convenient to use a lambda-derived phage as a source of dna, a proto ...	2001	18265266
evolution of the iss gene in escherichia coli.	the increased serum survival gene iss has long been recognized for its role in extraintestinal pathogenic escherichia coli (expec) virulence. iss has been identified as a distinguishing trait of avian expec but not of human expec. this gene has been localized to large virulence plasmids and shares strong similarities with the bor gene from bacteriophage lambda. here, we demonstrate that three alleles of iss occur among e. coli isolates that appear to have evolved from a common lambda bor precurs ...	2008	18281426
facile detection of specific rna-polypeptide interactions by maldi-tof mass spectrometry.	a simple method for the detection of specific rna-polypeptide interactions using maldi-tof mass spectroscopy is described. instead of direct observation of the rna-polypeptide complex, we attempted the indirect observation of the binding event by focusing on the disappearance of the free polypeptide signal upon interaction with rna. as a result, specific binding of the rev-response element (rre) rna of the hiv with two rre-binding peptide aptamers, dla and rla peptides, as well as the bacterioph ...	2008	18288633
quantitative predictions for dna two-dimensional display according to size and nucleotide sequence composition.	2-d dna display is a simple separation method that provides a fast and economical way of visualizing polymorphism and comparing genomes. the dna fragments are separated first according to their size by standard gel electrophoresis and then according to their sequence composition using denaturing gradient gel electrophoresis. first developed by fischer and lerman (cell 1979, 16, 191-200), this method has recently been used to distinguish strains within a bacterial species. the genomic restriction ...	2008	18288775
surface-directed and ethanol-induced dna condensation on mica.	the adsorption of lambda-phage dna onto mica was investigated with atomic force microscopy. we found that the morphologies depended on the solvent conditions in the sample preparation procedure. flat-lying networks of hybridized single-stranded dna were obtained if ultrapure water was used. if buffered conditions are maintained during the whole of the preparation procedure, single double-stranded dna molecules are adsorbed. the adsorbed double-stranded dna molecules subsequently can be condensed ...	2008	18293959
host responses influence on the induction of lambda prophage.	inactivation of bacteriophage lambda ci repressor leads almost exclusively to lytic development. prophage induction can be initiated either by dna damage or by heat treatment of a temperature-sensitive repressor. these two treatments also cause a concurrent activation of either the host sos or heat-shock stress responses respectively. we studied the effects of these two methods of induction on the lytic pathway by monitoring the activation of different lambda promoters, and found that the lambda ...	2008	18298445
mechanisms of specific and nonspecific binding of architectural proteins in prokaryotic gene regulation.	ihf and hu are small basic proteins of eubacteria that bind as homodimers to double-stranded dna and bend the duplex to promote architectures required for gene regulation. these architectural proteins share a common alpha/beta fold but exhibit different nucleic acid binding surfaces and distinct functional roles. with respect to dna-binding specificity, for example, ihf is sequence specific, while hu is not. we have employed raman difference spectroscopy and gel mobility assays to characterize t ...	2008	18302340
stochastic receptor expression allows sensitive bacteria to evade phage attack. part i: experiments.	it has long been suspected that population heterogeneity, either at a genetic level or at a protein level, can improve the fitness of an organism under a variety of environmental stresses. however, quantitative measurements to substantiate such a hypothesis turn out to be rather difficult and have rarely been performed. herein, we examine the effect of expression heterogeneity of lambda-phage receptors on the response of an escherichia coli population to attack by a high concentration of lambda- ...	2008	18310238
evaluation of t7 and lambda phage display systems for survey of autoantibody profiles in cancer patients.	in the current study we attempted to evaluate the suitability of t7 select 10-3b and lambdakm8 phage display systems for the identification of antigens eliciting b cell responses in cancer patients and the production of phage-displayed antigen microarrays that could be exploited for the monitoring of autoantibody profiles. members of 15 tumour-associated antigen (taa) families were cloned into both phage display vectors and the taa mini-libraries were immunoscreened with 22 melanoma patients' se ...	2008	18314130
a biotin interference assay highlights two different asymmetric interaction profiles for lambda integrase arm-type binding sites in integrative versus excisive recombination.	the site-specific recombinase integrase encoded by bacteriophage lambda promotes integration and excision of the viral chromosome into and out of its escherichia coli host chromosome through a holliday junction recombination intermediate. this intermediate contains an integrase tetramer bound via its catalytic carboxyl-terminal domains to the four "core-type" sites of the holliday junction dna and via its amino-terminal domains to distal "arm-type" sites. the two classes of integrase binding sit ...	2008	18319248
involvement of dna replication in phage lambda red-mediated homologous recombination.	crosses between a non-replicating linear bacteriophage lambda chromosome and a replicating plasmid bearing a short cloned segment of lambda dna were monitored by extracting dna from infected cells, and analysing it via restriction endonuclease digestion and southern blots. recombinant formation resulting from the action of the red homologous recombination system, observed directly in this way, was found to be fast, efficient, independent of the bacterial reca function and highly dependent upon r ...	2008	18333884
[gene cloning, selection of plasmids and application of fasciola hepatica cathepsin l1 gene].	gene cloning refers to the process by which a fragment of dna is transferred from one organism to a vector. a vector is an agent that can carry a dna fragment into a host cell. commonly used vectors include plasmids, lambda phage, cosmid and yeast artificial chromosome (yac). plasmid vectors that have been extensively used in genetic engineering are derived from natural plasmids. these contain a genetic marker conferring a phenotype that can be selected for or against and a polylinker or multipl ...	2008	18351545
extended function of plasmid partition genes: the sop system of linear phage-plasmid n15 facilitates late gene expression.	the mitotic stability of the linear plasmid-prophage n15 of escherichia coli depends on a partition system closely related to that of the f plasmid sopabc. the two sop systems are distinguished mainly by the arrangement of their centromeric sopb-binding sites, clustered in f (sopc) and dispersed in n15 (ir1 to ir4). because two of the n15 inverted repeat (ir) sites are located close to elements presumed (by analogy with phage lambda) to regulate late gene expression during the lytic growth of n1 ...	2008	18359814
bacteriophage infection is targeted to cellular poles.	the poles of bacteria exhibit several specialized functions related to the mobilization of dna and certain proteins. to monitor the infection of escherichia coli cells by light microscopy, we developed procedures for the tagging of mature bacteriophages with quantum dots. surprisingly, most of the infecting phages were found attached to the bacterial poles. this was true for a number of temperate and virulent phages of e. coli that use widely different receptors and for phages infecting yersinia ...	2008	18363799
identification and functional analysis of the rz/rz1-like accessory lysis genes in the membrane-containing bacteriophage prd1.	bacteriophage prd1 is a tailless membrane-containing double-stranded (ds) dna virus infecting a variety of gram-negative bacteria. in order to affect cell lysis, like most dsdna phages, prd1 uses the holin-endolysin system. in this study, we identified two accessory lysis genes, xxxvi and xxxvii, coding for proteins p36 and p37, respectively. using genetic complementation assays, we show that protein pair p36/p37 is a functional and interchangeable analogue of the rz/rz1 of bacteriophage lambda. ...	2008	18366440
znf397, a new class of interphase to early prophase-specific, scan-zinc-finger, mammalian centromere protein.	the centromere is a complex structure required for equal segregation of newly synthesised sister chromatids at mitosis. one of the significant objectives in centromere research is to determine the complete repertoire of protein components that constitute the kinetochore. here, we identify a novel centromere protein using a centromere-positive autoimmune serum from a patient with watermelon stomach disease. western blot and screening of a lambda phage expression library revealed a 60-kda protein, ...	2008	18369653
dna looping can enhance lysogenic ci transcription in phage lambda.	the lysogenic state of bacteriophage lambda is maintained by ci repressor, which negatively regulates two promoters to block lytic gene expression. expression of ci is itself controlled by positive and negative feedback as ci binds to o(r) to regulate the p(rm) promoter. in addition to direct interactions with operator dna, ci tetramers bound at o(l) and o(r) can come together to form an octamer, looping the dna that lies between them and allowing o(l) to assist with negative regulation of p(rm) ...	2008	18391225
binding cooperativity in phage lambda is not sufficient to produce an effective switch.	in the wild-type phage lambda, binding of ci to o(r)2 helps polymerase bound to p(rm) transition from a closed to open complex. activators on other promoters increase the polymerase-dna binding energy, or affect both the binding energy and the closed-open transition probability. using a validated mathematical model, we show that these two modes of upregulation have very different effects on the promoter function. we predict that if ci(2) bound to o(r)2 produced equal increase in rnap-dna binding ...	2008	18400951
expression cloning of neural genes using xenopus laevis oocytes.	expression cloning requires a representative cdna or genomic dna library and a host organism in which the cloned genes can be transcribed and/or translated. it likewise requires a method to detect the expressed protein using, for example, the inherent biological activity of the gene or antibodies specific for the gene product. most successful expression cloning strategies have employed cdna libraries constructed in plasmid or bacteriophage lambda vectors and xenopus oocytes or cultured mammalian ...	2001	18428487
crystal structure of the lambda repressor and a model for pairwise cooperative operator binding.	bacteriophage lambda has for many years been a model system for understanding mechanisms of gene regulation. a 'genetic switch' enables the phage to transition from lysogenic growth to lytic development when triggered by specific environmental conditions. the key component of the switch is the ci repressor, which binds to two sets of three operator sites on the lambda chromosome that are separated by about 2,400 base pairs (bp). a hallmark of the lambda system is the pairwise cooperativity of re ...	2008	18432246
genotoxicity of wastewaters used for irrigation of food crops.	in most towns of india, wastewater coming from both industrial and domestic sources and without any treatment is used to irrigate the agricultural crops. this practice has been polluting the soil, and pollutants could possibly reach the food chain. for the above reasons, the wastewaters of ghaziabad city (india), which is used for irrigation, were sampled (at two different sites) and monitored for the presence of genotoxic agents from january 2005 to june 2007. gas chromatographic analysis showe ...	2009	18442071
a quantitative competitive pcr method to determine the parasite load in the brain of toxoplasma gondii-infected mice.	efficacy of vaccine candidates against toxoplasmosis may be expressed in terms of reduction in cyst number in brains of animals vaccinated and then challenged with a cyst-forming strain of toxoplasma gondii, compared to non-vaccinated animals. cyst number generally has been determined by microscopic examination of brain homogenate samples, a technique which has a low sensitivity and is time-consuming. here we describe a quantitative competitive pcr method, which allows quantifying t. gondii dna ...	2008	18456545
[gene modification in the genome of epstein-barr virus cloned as a bacterial artificial chromosome].	epstein-barr virus (ebv) is an oncogenic herpesvirus associated with a variety of malignancies, including burkitt's lymphoma and nasopharyngeal carcinoma (npc). functions of most ebv genes have not been determined. the use of bacterial artificial chromosome (bac) to clone and modify the genome of ebv has enhanced the gene function study in the context of genome. infectious clones of ebv were previously established by using ebv-bac plasmid p2089. in order to further investigate ebv mutant biology ...	2008	18479068
expanding the realm of ultrafast protein folding: gpw, a midsize natural single-domain with alpha+beta topology that folds downhill.	all ultrafast folding proteins known to date are either very small in size (less than 45 residues), have an alpha-helix bundle topology, or have been artificially engineered. in fact, many of them share two or even all three features. here we show that gpw, a natural 62-residue alpha+beta protein expected to fold slowly in a two-state fashion, folds in microseconds (i.e., from tau = 33 micros at 310 k to tau = 1.7 micros at 355 k). thermodynamic analyses of gpw reveal probe dependent thermal den ...	2008	18479088
nongenetic individuality in the host-phage interaction.	isogenic bacteria can exhibit a range of phenotypes, even in homogeneous environmental conditions. such nongenetic individuality has been observed in a wide range of biological processes, including differentiation and stress response. a striking example is the heterogeneous response of bacteria to antibiotics, whereby a small fraction of drug-sensitive bacteria can persist under extensive antibiotic treatments. we have previously shown that persistent bacteria enter a phenotypic state, identifie ...	2008	18494559
rapid genome sequencing with short universal tiling probes.	the increasing availability of high-quality reference genomic sequences has created a demand for ways to survey the sequence differences present in individual genomes. here we describe a dna sequencing method based on hybridization of a universal panel of tiling probes. millions of shotgun fragments are amplified in situ and subjected to sequential hybridization with short fluorescent probes. long fragments of 200 bp facilitate unique placement even in large genomes. the sequencing chemistry is ...	2008	18500336
analysis of some phenotypic traits of feces-borne temperate lambdoid bacteriophages from different immunity groups: a high incidence of cor+, fhua-dependent phages.	a group of previously isolated heterogeneous mep lambdoid phages (43) from 19 different immunity groups for phage infection was further characterized to gain insight into some phenotypic traits and to assess their relationship with phage lambda. interestingly, the fhua host receptor was required by the majority of mep phages (37 out of 43; approximately 85%). the cor gene, which has been reported to be involved in fhua-dependent exclusion of lambdoid phages, was also found in most of the fhua-de ...	2008	18516490
prophages in marine bacteria: dangerous molecular time bombs or the key to survival in the seas?	bacteriophages are realized to be numerous and important components of oceanic food webs principally because of their lytic capabilities. the subtle changes that temperate phages impart to their hosts in the oceans are far less understood. occurrences of lysogeny in the oceans correlate well with conditions unfavorable for rapid host growth. in coliphage lambda, phage encoded repressors have been shown to modulate host metabolic gene expression and phenotype, resulting in economizing host energy ...	2008	18521076
fluorescence resonance energy transfer sensors for quantitative monitoring of pentose and disaccharide accumulation in bacteria.	engineering microorganisms to improve metabolite flux requires detailed knowledge of the concentrations and flux rates of metabolites and metabolic intermediates in vivo. fluorescence resonance energy transfer sensors represent a promising technology for measuring metabolite levels and corresponding rate changes in live cells. these sensors have been applied successfully in mammalian and plant cells but potentially could also be used to monitor steady-state levels of metabolites in microorganism ...	2008	18522753
evolutionary dominance of holin lysis systems derives from superior genetic malleability.	for the microviruses and the leviviruses, bacteriophages with small single-stranded genomes, host lysis is accomplished by expression of a single gene that encodes an inhibitor of cell wall synthesis. in contrast, phages with double-stranded dna genomes use a more complex system involving, at minimum, an endolysin, which degrades peptidoglycan, and a holin, which permeabilizes the membrane in a temporally programmed manner. to explore the basis of this difference, a chimera was created in which ...	2008	18524925
crystallization and structure determination of the core-binding domain of bacteriophage lambda integrase.	bacteriophage lambda integrase catalyzes site-specific dna recombination. a helical bundle domain in the enzyme, called the core-binding domain (int(cb)), promotes the catalysis of an intermediate dna-cleavage reaction that is critical for recombination and is not well folded in solution in the absence of dna. to gain structural insights into the mechanism behind the accessory role of this domain in catalysis, an attempt was made to crystallize an int(cb)-dna complex, but crystals of free int(cb ...	2008	18540053
preparation of multimilligram quantities of large, linear dna molecules for structural studies.	we describe a method for preparing large, linear dna molecules in amounts that are suitable for structural studies. the procedure employs self-primed dna amplification on a starting molecule that consists of the sequence of interest flanked by the cohesive end sequences from bacteriophage lambda as well as endonuclease recognition sites. amplification produces long polymers of dna, tens of kilobases in length, which harbor many copies of the sequence of interest. endonuclease digestion of these ...	2008	18547516