Publications

expression of the escherichia coli ftsz gene: trials and tribulations of gene fusion studies.	the ftsz gene of escherichia coli, which codes for an essential cell division protein, is subjected to multiple regulation, as shown in part with studies using an ftsz::lacz operon fusion located on phage lambda jfl100. using this same fusion, we sought to isolate regulatory mutants overexpressing ftsz by selecting mutants able to grow on lactose. one lac+ mutant was obtained which overexpressed the ftsz::lacz fusion 70-fold. the mutation responsible for the overexpression lies in a new gene, co ...	1993	8468005
from the double-helix to novel approaches to the sequencing of large genomes.	elucidation of the structure of dna by watson and crick [nature 171 (1953) 737-738] has led to many crucial molecular experiments, including studies on dna replication, transcription, physical mapping, and most recently to serious attempts directed toward the sequencing of large genomes [watson, science 248 (1990) 44-49]. i am totally convinced of the great importance of the human genome project, and toward achieving this goal i strongly favor 'top-down' approaches consisting of the physical map ...	1993	8276270
the mode of replication is a major factor in segregational plasmid instability in lactococcus lactis.	the effects of the rolling-circle and theta modes of replication on the maintenance of recombinant plasmids in lactococcus lactis were studied. heterologous escherichia coli or bacteriophage lambda dna fragments of various sizes were inserted into vectors based on either the rolling-circle-type plasmid pwv01 or the theta-type plasmid pambeta1. all pambeta1 derivatives were stably maintained. pwv01 derivatives, however, showed size-dependent segregational instability, in particular when large dna ...	1993	16348863
recognition of boxa antiterminator rna by the e. coli antitermination factors nusb and ribosomal protein s10.	the boxa sequences of the e. coli ribosomal rna (rrn) operons are sufficient to cause rna polymerase to read through rho-dependent transcriptional terminators. we show that a complex of the transcription antitermination factors nusb and ribosomal protein s10 interacts specifically with boxa rna. neither nusb nor s10 binds boxa rna on its own, and neither nusa nor nusg affects the interaction of the nusb-s10 complex with boxa rna. mutations in boxa that impair its antitermination activity comprom ...	1993	7678781
characterization of gene expression in recombinant escherichia coli cells infected with phage lambda.	phage lambda infection was investigated for possible production of toxic foreign proteins in escherichia coli. the target gene transcription was regulated by a t7 promoter, which was initiated under the action of t7 rna polymerase delivered by infecting phage. two types of phage infection were investigated. in both cases, deletion of the int gene prevents lysogenic integration. one phage, lambda ce6, contains the sam7 lysis mutation, so that infectious phage particles remain intracellular. the o ...	1993	7763591
in vivo regulatory responses of four escherichia coli operons which encode leucyl-trnas.	four escherichia coli operons, the leuv operon which encodes trna(1leu), the leux operon which encodes trna(6leu), the mett operon which encodes trna(3leu), and the argt operon which encodes trna(1leu), were examined for the stringent response induced by serine hydroxamate and for growth rate-dependent regulation. in nuclease protection assays, the leuv operon displayed the stringent response in response to leucine starvation, analog inhibition, and growth of a temperature-sensitive leucyl-trna ...	1993	7680341
spontaneous transposition in the bacteriophage lambda cro gene residing on a plasmid.	a new mutagenesis assay system based on the phage lambda cro repressor gene residing on a plasmid was developed. the assay detects mutations in cro that decrease the binding of the repressor to the or operator in an or pr-lacz fusion present in a lambda prophage. mutations arose spontaneously during growth of e. coli cells harboring cro plasmids at a frequency of 3-6 x 10(-6). analysis of some 200 cro mutants from several 'wild-type' strains revealed a substantial fraction of 25-70% insertion ev ...	1992	1373849
both forms of translational initiation factor if2 (alpha and beta) are required for maximal growth of escherichia coli. evidence for two translational initiation codons for if2 beta.	the gene infb codes for two forms of translational initiation factor if2; if2 alpha (97,300 da) and if2 beta (79,700 da). if2 beta arises from an independent translational event on a gug codon located 471 bases downstream from if2 alpha start codon. by site-directed mutagenesis we constructed six different mutations of this gug codon. in all cases, if2 beta synthesis was variably affected by the mutations but not abolished. we show that the residual expression of if2 beta results from translatio ...	1992	1374802
characterization of the transcription activator protein c1 of bacteriophage p22.	we cloned, expressed, and purified the positive regulatory protein c1 of the temperate phage p22 of salmonella typhimurium. the purified protein was characterized as to its amino acid composition, protein sequence, molecular weight, and antigenicity. p22 c1 was shown to be a tetrameric protein composed of four identical subunits with m(r) = 10,000. moreover, we identified and characterized two p22 c1-dependent phage promoters, p(re) and pa23, whose function was completely dependent on c1 both in ...	1992	1385814
construction of coliphage lambda charon vectors with bamh1 cloning sites. 1980.		1992	1422018
overproduction and purification of bacillus subtilis dna polymerase iii.	the objectives of this work were to engineer the cloned polc gene encoding bacillus subtilis dna polymerase iii for controlled overexpression in escherichia coli and to devise a facile purification scheme permitting the large-scale production of pure recombinant polymerase. the translational signals of polc were restructured by expression cassette pcr (macferrin et al., 1990, proc. natl. acad. sci. usa 87, 1937-1941), and the modified gene was inserted into the expression plasmid, pkc30 (rosenbe ...	1992	1422209
the translation initiation site of recombinant trypanosoma brucei ornithine decarboxylase varies with different promoters.	expression of the trypanosoma brucei ornithine decarboxylase (odc) gene in escherichia coli behind the lambda phage pr promoter led to the production of a recombinant enzyme having the same subunit molecular weight as the native enzyme [4]. however, when the same gene is expressed behind the tac promoter or the phoa promoter, the odcs produced by the transformed e. coli have subunit molecular weights approximately 2 kda higher than that of the native enzyme. amino terminal sequencing of the reco ...	1992	1435879
bacteriophage lambda papa: not the mother of all lambda phages.	the common laboratory strain of bacteriophage lambda--lambda wild type or lambda papa--carries a frameshift mutation relative to ur-lambda, the original isolate. the ur-lambda virions have thin, jointed tail fibers that are absent from lambda wild type. two novel proteins of ur-lambda constitute the fibers: the product of stf, the gene that is disrupted in lambda wild type by the frameshift mutation, and the product of gene tfa, a protein that is implicated in facilitating tail fiber assembly. r ...	1992	1439823
involvement of the escherichia coli rna polymerase alpha subunit in transcriptional activation by the bacteriophage lambda ci and cii proteins.	escherichia coli cells harbouring the rpoa341 mutation produce an rna polymerase which transcribes inefficiently certain operons subject to positive control. here, we demonstrate that the rpoa341 allele also prevents lysogenization of the host strain by bacteriophage lambda, a process dependent upon the action of two phage-encoded activators. this phenomenon was shown to arise from an inability to establish an integrated prophage rather than a failure to maintain the lysogenic state. the inabili ...	1992	1452017
enriched sources of escherichia coli replication proteins. the dnag primase is a zinc metalloprotein.	primase, the product of the escherichia coli dnag gene, is the enzyme responsible for rna primer synthesis on both template strands at replication forks during chromosomal dna synthesis. the dnag gene was modified by replacement of the natural ribosome-binding site with one complementary to the 3' end of 16s rrna, and then inserted downstream of tandem bacteriophage lambda pr and pl promoters in the puc9-derived vector pce30. following thermal induction of transcription, the resulting plasmid pp ...	1992	1511009
effect of escherichia coli nusg function on lambda n-mediated transcription antitermination.	the escherichia coli nus factors act in conjunction with the bacteriophage lambda n protein to suppress transcription termination on the lambda chromosome. nusa binds both n and rna polymerase and may also interact with other nus factors. to search for additional components of the n antitermination system, we isolated host revertants that restored n activity in nusa1 mutants. one revertant, nusg4, was mapped to the rif region of the e. coli chromosome and shown to represent a point mutation near ...	1992	1531224
lambda int protein bridges between higher order complexes at two distant chromosomal loci attl and attr.	the excisive recombination reaction of bacteriophage lambda involves a specific and efficient juxtaposition of two distant higher order protein-dna complexes on the chromosome of escherichia coli. these complexes, which mediate synapsis and strand exchange, consist of two dna sequences, attl and attr, the bivalent dna binding protein int, and the sequence-specific dna bending proteins, ihf, xis, and fis. the protein-protein and protein-dna interactions within, and between, these complexes were s ...	1992	1533056
an efficient phage plaque screen for the random mutational analysis of the interaction of hiv-1 gp120 with human cd4.	a lambda phage expression methodology was adapted to dissect protein/ligand interactions efficiently through the creation and rapid screening of large numbers of mutants. here we describe the method and its specific application to the interaction between the external envelope glycoprotein of the human immunodeficiency virus (hiv-1), gp120, and the human cell surface protein cd4. random substitutions were introduced throughout the gp120 binding region (amino acids 38-62) in the amino-terminal dom ...	1992	1533631
t41 mutation in lac repressor is tyr282----asp.	a monomeric mutant of the lac repressor protein, designated t41, produced originally by the in vivo mutt mutagenesis exhibited behavior similar to mutants identified as tyr282----ser or tyr282----gln [schmitz et al., j. biol. chem. 251 (1976) 3359-3366]. the t41 gene, encoded within a phage lambda prophage in the escherichia coli genome, was amplified by polymerase chain reaction and sequenced. the only mutation found in the nucleotide sequence corresponded to position 282 in the protein, and th ...	1992	1547952
cloning and nucleotide sequence of the escherichia coli cytidine deaminase (ccd) gene.	the structural gene that encodes cytidine deaminase (cdd) in escherichia coli was cloned from kohara phage lambda 365 (7f1), and its nucleotide sequence was determined. plasmids harboring the gene complemented chromosomal cdd mutations, enhanced cytidine deaminase activity in cell extracts, and directed the synthesis of a protein identical in mass and n-terminal amino acid sequence with cytidine deaminase purified from wild-type bacteria. metal analysis of the purified, plasmid-encoded deaminase ...	1992	1567863
cloning and overexpression of the lactobacillus bulgaricus nad(+)-dependent d-lactate dehydrogenase gene in escherichia coli:purification and characterization of the recombinant enzyme.	the lactobacillus bulgaricus nad(+)-dependent d-lactate dehydrogenase gene was amplified by the polymerase chain reaction and cloned into an escherichia coli expression plasmid pkk223.3. attempts to clone the full-length chromosomal dna encoding d-lactate dehydrogenase from a partial sau3ai lambda phage library or an enriched clone bank in e. coli were unsuccessful. the recombinant plasmid pkbuldh containing the amplified gene overexpressed d-lactate dehydrogenase (greater than 30% of total solu ...	1992	1610363
renaturation of cobra venom phospholipase a2 expressed from a synthetic gene in escherichia coli.	cobra venom (naja naja naja) phospholipase a2 (pla2) contains 14 cysteines in the form of 7 disulfide bonds amongst its 119 amino acids. a gene encoding the pla2 was synthesized and inserted into a bacterial expression vector containing the phage lambda pl promoter. in order to obtain protein without the initiating methionine at the n-terminus, a factor xa site was engineered upstream from the pla2 gene. upon heat-induction of the cells transformed with the expression plasmid, the protein is pro ...	1992	1730025
saccharomyces cerevisiae elongation factor 2. genetic cloning, characterization of expression, and g-domain modeling.	the elongation factor 2 (ef-2) genes of the yeast saccharomyces cerevisiae have been cloned and characterized with the ultimate goal of gaining a better understanding of the mechanism and control of protein synthesis. two genes (eft1 and eft2) were isolated by screening a bacteriophage lambda yeast genomic dna library with an oligonucleotide probe complementary to the domain of ef-2 that contains diphthamide, the unique posttranslationally modified histidine that is specifically adp-ribosylated ...	1992	1730643
phage lambda has an analog of escherichia coli reco, recr and recf genes.	the recf pathway catalyzes generalized recombination in escherichia coli that is mutant for recbc, sbcb and sbcc. this pathway operating on conjugational recombination requires the reca, recf, recj, recn, reco, recq, recr, ruva, ruvb and ruvc genes. in contrast, lambda mutant for its own recombination genes, int, red alpha and red beta, requires only the reca and recj genes to recombine efficiently in recbc sbcb sbcc cells. deletion of an open reading frame in the ninr region of lambda results i ...	1992	1310087
alleviation of ecok dna restriction in escherichia coli and involvement of umudc activity.	the activity of the ecok dna restriction system of escherichia coli reduces both the plating efficiency of unmodified phage lambda and the transforming ability of unmodified pbr322 plasmid dna. however, restriction can be alleviated in wild-type cells, by uv irradiation and expression of the sos response, so that 10(3)- to 10(4)-fold increases in phage growth and fourfold increases in plasmid transformation occurred with unmodified dna. restriction alleviation was found to be a transient effect ...	1992	1310522
nonrandom orientation of transposon tn5supf insertions in phage lambda.	transposition of mini-transposon tn5supf to phage lambda can be selected in two ways: (i) by plaque formation on a dnab amber strain of escherichia coli, which requires expression of the transposon-borne suppressor trna gene (supf) during lytic phage growth, or (ii) by lysogenization of a strain with amber mutations in tet and amp resistance genes, and selection of tcr apr (sup+) transductant colonies. tn5supf insertions in several lambda clones were isolated and mapped using a polymerase chain ...	1992	1316868
the construction of streptomyces cyaneus genomic libraries in escherichia coli is dependent upon the use of mcr-deficient strains.	streptomyces cyaneus genomic dna ligated into either lambda phage or plasmid vectors was very inefficiently cloned into standard escherichia coli host strains. however, the same material could be efficiently cloned using mcr-deficient e. coli strains. these results suggest that the s. cyaneus genome contains 5-methylcytosine residues, some of which occur within the recognition sequences of the e. coli mcr restriction system.	1992	1327960
a selective lambda phage cloning vector with automatic excision of the insert in a plasmid.	a bacteriophage lambda cloning vehicle has been constructed for the generation of cdna libraries. the vector has the following properties. (1) it has a unique bamhi site engineered into the lambda gam gene. segments of dna can be cloned into this site and clones with an insert can be selected by their ability to grow on an escherichia coli host lysogenic for phage p2 (spi- phenotype). (2) when the recombinant phage infects a cre-producing e. coli strain, a site-specific recombination event resul ...	1992	1327972
efficient large-scale sequencing of the escherichia coli genome: implementation of a transposon- and pcr-based strategy for the analysis of ordered lambda phage clones.	we have developed a strategy for efficient sequence analysis of the genome of e. coli k-12 using insertions of a tn5-derived mini-transposon into overlapping ordered lambda phage clones to provide universal primer-binding sites, and pcr amplification of dna segments adjacent to the insertions. transposon-containing clones were selected by blue plaque formation on a dnabamber laczamber e. coli strain. insertion points every 0.5-1 kb were identified by 'analytical pcr' and segments between the tra ...	1992	1336178
construction and characterization of a single-chain catalytic antibody.	the antigen-binding (fab) fragment of the catalytic monoclonal antibody npn43c9 has recently been cloned by using bacteriophage lambda. by inserting the variable regions of this fab coding sequence into a (nh2)-vl-linker-vh-(cooh) construct (where vl and vh represent the heavy and light chain variable regions), we have assembled a recombinant gene encoding a catalytic single-chain antigen-binding protein. this protein has been expressed in escherichia coli and exhibits the same catalytic paramet ...	1991	2023948
structural and functional comparison between the stability systems pard of plasmid r1 and ccd of plasmid f.	the stability determined by the systems pard of plasmid r1 and ccd of plasmid f is due to the concerted action of two proteins, a cytotoxin and an antagonist of this function. in this paper we report that ccda and kis proteins, the antagonists of the ccd and pard systems respectively, share significant sequence homologies at both ends. in kis, these regions seem to correspond to two different domains. despite the structural similarities, kis and ccda are not interchangeable. in addition we have ...	1991	2017133
cloning, sequencing and overexpression of the gene for prokaryotic factor ef-p involved in peptide bond synthesis.	a soluble protein ef-p (elongation factor p) from escherichia coli has been purified and shown to stimulate efficient translation and peptide-bond synthesis on native or reconstituted 70s ribosomes in vitro. based on the partial amino acid sequence of ef-p, 18- and 24-nucleotide dna probes were synthesized and used to screen lambda phage clones from the kohara gene bank. the entire ef-p gene was detected on lambda clone #650 which contains sequences from the 94 minute region of the e.coli genome ...	1991	1956781
rna polymerases from pseudomonas aeruginosa and pseudomonas syringae respond to escherichia coli activator proteins.	the activities of rna polymerases (rnaps) from pseudomonas aeruginosa and pseudomonas syringae were compared with that of escherichia coli rnap. all three enzymes are able to initiate transcription at the trpba promoter of p. aeruginosa and at the coliphage lambda promoters, prm and pre, in response to heterospecific activators (trpi protein, repressor, and cii protein, respectively). however, both pseudomonas polymerases have less stringent requirements for promoter recognition in the absence o ...	1991	1898924
selection of lacz operon fusions in genes of gluconate metabolism in e. coli. characterization of a gntt::lacz fusion.	the initial steps involved in the utilization of gluconate by e. coli, its incorporation into the cell and subsequent phosphorylation to gluconate 6-phosphate, conform two systems that duplicate activities. these systems, gnti and gntii, are specified by two sets of genes distinctly regulated and located respectively at the mala-asd (75 min) and fdp-vals (96 min) regions of the bacterial chromosome. the presence of duplicate activities in the metabolism of gluconate of e. coli, has made difficul ...	1991	1843569
recombinagenic processing of uv-light photoproducts in nonreplicating phage dna by the escherichia coli methyl-directed mismatch repair system.	nonreplicating lambda phage dna in homoimmune escherichia coli lysogens provides a useful model system for study of processes that activate dna for homologous recombination. we measured recombination by extracting phage dna from infected cells, using it to transfect reca recipient cells, and scoring the frequency of recombinant infective centers. with unirradiated phage, recombinant frequencies were less than 0.1%. however, recombination could be increased over 300-fold by prior uv irradiation o ...	1991	1838344
bacteriophage lambda promoters pl and pr: sequence determinants of in vivo activity and of sensitivity to the dna gyrase inhibitor, coumermycin.	sequence encompassing the region between bp -43 and +8 of the pl and pr promoters of bacteriophage lambda, as well as sequence variants of these promoters, were compared with respect to their ability to drive a promoterless cat gene in vivo. for both pl- and pr-based promoters, variants with one nonconsensus bp rather than the consensus promoter were found to be maximally active. determination of promoter function in csh26 and c600 revealed a marked strain dependence in the activity of some prom ...	1991	1847348
hu and integration host factor function as auxiliary proteins in cleavage of phage lambda cohesive ends by terminase.	hu and integration host factor (ihf) are small, basic heterodimeric dna-binding proteins which participate in transcription initiation, dna replication, and recombination. we constructed isogenic escherichia coli strains in which hu, ihf, or both proteins were absent. bacteriophage lambda did not grow in hosts lacking both hu and ihf. phage dna replication and late gene transcription were normal in the double mutants, but packaging of lambda dna was defective. mature phage dna molecules were abs ...	1991	1825651
isolation and preliminary characterization of escherichia coli mutants resistant to lethal action of the bacteriophage lambda p gene.	both spontaneous and ntg-induced mutants of escherichia coli 594 insensitive to the lethal action of lambda p gene were isolated and called rpl (resistant to p lethality). these mutants were of two types, showing different phenotypes. on type i rpl mutants, lambda cl- and lambda v1v3 did not plate, while lambda vir, lambda cl- c17, lambda imm434, and lambda imm21 did; plasmid pmr45 carrying the lambda p gene could not complement lambda imm21p- phage in type i mutants. on the other hand, the type ...	1991	1827225
identification of protein binding sites in genomic dna by two-dimensional gel electrophoresis.	we describe a simple two-dimensional electrophoresis procedure to identify the recognition sites of dna-binding proteins within large dna molecules. using this approach, we have mapped e. coli ihf (integration host factor) binding sites within phage lambda (48 kb) and phage mu (39 kb) dna. we are also able to visualize ihf binding sites in e. coli chromosomal dna (4,700 kb). we present an extension of this technique using direct amplification by pcr of the isolated restriction fragments, which s ...	1991	1827523
heat-inducible reactivation of uv-damaged bacteriophage lambda.	induction of the sos response in uv-irradiated bacteria leads to an increase in the survival of an infecting irradiated bacteriophage lambda (weigle 1953). we report that a similar reactivation of irradiated phage lambda was induced by shifting the culture of recipient bacteria from 30 degrees to 47 degrees c. however, this repair process was nonmutagenic. the amplitude of the phenomenon was increased with the quantity of uv lesions in the phage dna. it was present despite mutations affecting th ...	1991	1827875
dna recognition by the helix-turn-helix motif: investigation by laser raman spectroscopy of the phage lambda repressor and its interaction with operator sites ol1 and or3.	the lambda repressor provides a model system for biophysical studies of dna recognition by the helix-turn-helix motif. we describe laser raman studies of the lambda operator sites ol1 and or3 and their interaction with the dna-binding domain of lambda repressor (residues 1-102). raman spectra of the two dna sites exhibit significant differences attributable to interstrand purine-purine steps that differ in the two oligonucleotides. remarkably, the conformation of each operator is significantly a ...	1991	1828373
heteroduplex chain polarity in recombination of phage lambda by the red, recbcd, recbc(d-) and recf pathways.	we have examined the chain polarity of heteroduplex dna in unreplicated, bacteriophage lambda splice recombinants when recombination was by the recbcd, recbc(d-), or recf pathway of escherichia coli or the red pathway of lambda. for each of these pathways, recombination is activated by the cutting of cos that accompanies chromosome packaging, and is effected by recombination enzymes acting at the right end created by that cutting. for exchanges occurring near cos, one parent makes a lesser physi ...	1991	1829428
efficient excision of phage lambda from the escherichia coli chromosome requires the fis protein.	the escherichia coli protein fis has been shown to bind a single site in the recombination region of phage lambda and to stimulate excisive recombination in vitro (j. f. thompson, l. moitoso de vargas, c. koch, r. kahmann, and a. landy, cell 50:901-908, 1987). we demonstrate that mutant strains deficient in fis expression show dramatically reduced rates of lambda excision in vivo. phage yields after induction of a stable lysogen are reduced more than 200-fold in fis cells. the defect observed in ...	1991	1829453
the last duplex base-pair of the phage lambda chromosome. involvement in packaging, ejection and routing of lambda dna.	cosn is the site at which the bacteriophage lambda dna packaging enzyme, terminase, introduces staggered nicks to generate the cohesive ends of mature lambda chromosomes. genetic and molecular studies show that cosn is recognized specifically by terminase and that effects of cosn mutations on lambda dna packaging and cosn cleavage are well correlated. mutations affecting a particular base-pair of cosn are unusual in being lethal in spite of causing only a moderate defect in cosn cleavage and dna ...	1991	1830344
isolation and characterization of mutations in the bacteriophage lambda terminase genes.	the terminase enzyme of bacteriophage lambda is a hetero-oligomeric protein which catalyzes the site-specific endonucleolytic cleavage of lambda dna and its packaging into phage proheads; it is composed of the products of the lambda nul and a genes. we have developed a simple method to select mutations in the terminase genes carried on a high-copy-number plasmid, based on the ability of wild-type terminase to kill reca strains of escherichia coli. sixty-three different spontaneous mutations and ...	1991	1830578
molecular cloning and nucleotide sequence of the rhizobium phaseoli reca gene.	a recombinant lambda phage carrying the reca gene of rhizobium phaseoli was isolated from a r. phaseoli genomic library by complementation of the fec- phenotype of the recombinant phage in escherichia coli. when expressed in e. coli, the cloned reca gene was shown to restore resistance to both uv irradiation and the dna alkylating agent methyl methanesulphonate (mms). the r. phaseoli reca gene also promoted homologous recombination in e. coli. the cloned reca gene was only weakly inducible in e. ...	1991	1832737
dna sequence changes in mutations induced by ultraviolet light in the gpt gene on the chromosome of escherichia coli uvr+ and urva cells.	sequence changes in mutations induced by ultraviolet light are reported for the chromosomal escherichia coli gpt gene in almost isogenic e. coli uvr+ and excision-deficient uvra cells. differences between the mutagenic spectra are ascribed to preferential removal of photoproducts in the transcribed strand by excision repair in uvr+ cells. this conclusion is confirmed by analysis of published results for genes in both uvr+ and uvr- cells, showing a similar selective removal of mutagenic products ...	1991	1836051
[synthesis, secretion, and proteolytic degradation of diphtheria toxin in escherichia coli].	the recombinant plasmids have been constructed encoding the synthesis of a full-sized diphtheria toxin from its own or pr, pl-promoters of bacteriophage lambda in escherichia coli cells. the high level constitutive synthesis of toxin results in slow cell growth and plasmid elimination. the toxin was mainly detected in the periplasm, partially in the membrane and to a less extent in the cytoplasm and culturing medium. the dimeric form of toxin was found in the cytoplasm. participation of toxin b- ...	1991	1836836
the kila operon of promiscuous plasmid rk2: the use of a transducing phage (lambda pklaa-1) to determine the effects of the lethal klaa gene on escherichia coli cells.	the kil-kor regulon of promiscuous plasmid rk2 includes the replication initiator gene trfa and several potentially host-lethal kil loci (kila, kilb, kilc, kile), whose functions may be involved in plasmid maintenance or broad host range. the kila locus consists of a single operon of three genes (klaa, klab, klac), each of which is lethal when expressed from the klaa promoter in the absence of repressors encoded by kora and korb. in this study, we examined the effects of the unregulated klaa gen ...	1991	1838127
cyclic amp inhibits and putrescine represses expression of the spea gene encoding biosynthetic arginine decarboxylase in escherichia coli.	the spea gene of escherichia coli encodes biosynthetic arginine decarboxylase (adc), the first of two enzymes in a putrescine biosynthetic pathway. the activity of adc is negatively regulated by mechanisms requiring cyclic amp (camp) and camp receptor protein (crp) or putrescine. a 2.1-kb bamhi fragment containing the spea-metk intergenic region, spea promoter, and 1,389 bp of the 5' end of the spea coding sequence was used to construct transcriptional and translational spea-lacz fusion plasmids ...	1991	1646785
a combination of rnase h (rnh) and recbcd or sbcb mutations in escherichia coli k12 adversely affects growth.	colony forming ability of escherichia coli strains carrying the rnh-339::cat mutant allele is strongly dependent on the recbcd and sbcb genes. a mutation inactivating either the recbcd nuclease or exonuclease i (sbcb) is sufficient to restrict severely the efficiency of plating of strains carrying the rnh-339::cat mutation. combining a non-lethal temperature-sensitive mutation in the recbcd nuclease, recb270 (ts) or recc271 (ts), with rnh-339::cat renders strains temperature sensitive for growth ...	1991	1650908
effect of rfah (sfrb) and temperature on expression of rfa genes of escherichia coli k-12.	in order to study the regulation of a large block of contiguous genes at the rfa locus of escherichia coli k-12 which are involved in synthesis and modification of the lipopolysaccharide core, the transposon tnlacz was used to generate in-frame lacz fusions to the coding regions of five genes (rfaq, -g, -p, -b and -j) within this block. the beta-galactosidase activity of strains in which these fusions had been crossed into the chromosomal rfa locus was significantly decreased when the rfah11 (sf ...	1991	1655711
immunoanalysis of ultraviolet radiation induced dna damage and repair within specific gene segments of plasmid dna.	the region-specific heterogeneity of repairing dna damage has been established in several biological systems. a flexible and sensitive approach, based upon dna damage specific antibodies, is described to monitor the repair of specific lesions within discrete genomic segments. membrane transblotted dna restriction fragments are immunoanalyzed for the initial formation and repair of 254 nm radiation induced pyrimidine dimers. sensitivity of dimer immunodetection increases proportional to fragment ...	1991	1657185
reca-independent resistance to irradiation with u.v. light in acid-habituated escherichia coli.	growth of escherichia coli 1829 colv, i-k94 at ph 5.0 led to an increase in u.v. resistance compared with cells grown at ph 7.0. this was due to a phenotypic change, since organisms grown at ph 7.0 showed increased resistance after only 2.5-5.0 min incubation at the mildly acid ph. other e. coli k12 derivatives became more u.v.-resistant at ph 5.0 including uvra, reca and pola1 mutants. organisms grown at ph 5.0 also showed increased weigle reactivation of u.v.-irradiated lambda phage and this a ...	1991	2019551
transcription-dependent competition for a host factor: the function and optimal sequence of the phage lambda boxa transcription antitermination signal.	ordered development of lambdoid phages relies on systems of transcription termination and antitermination. the phage-encoded n early regulatory proteins, acting with the nus proteins of escherichia coli, modify rna polymerase to a form that overrides many transcription termination signals. these modifications require cis-acting sites, nut, located downstream of the early phage promoters. the nut sites in phages lambda, 21, and p22, which share similarities but are not identical, contain two sign ...	1990	2148536
a cdna encoding 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase from solanum tuberosum l.	a cdna encoding potato (solanum tuberosum l.) 3-deoxy-d-arabino-heptulosonate 7-phosphate synthase, the first enzyme of the shikimate pathway, was cloned into phage lambda gt11. the clone represents the first cdna for this enzyme from any eukaryotic source. the nucleotide sequence of the cdna was determined, and its identity was confirmed through partial amino acid sequence analysis of the encoded enzyme. the cdna contains a 1527-base pair open reading frame that encodes a polypeptide with a cal ...	1990	1967256
bacterial expression of immunoglobulin vh proteins.	a bacterial expression system in escherichia coli has been developed that produces as much as 10 mg/l of culture of the vh protein associated with monoclonal antibodies specific for the 5-dimethylaminonaphthalene-2-sulfonyl (dns) group. this system has been applied to the expression of the vh genes derived from a low-affinity, igm-producing hybridoma and from a high-affinity, igg-producing cell line. the plasmid vectors (contributed by dr william f. studier) utilize a t7 expression cassette whos ...	1990	2107393
effects of medium quality on the expression of human interleukin-2 at high cell density in fermentor cultures of escherichia coli k-12.	we examined the ability of transformed escherichia coli cells in fermentor cultures to accumulate interleukin-2 (il-2) intracellularly under temperature-regulated control of the phage lambda pl promoter. induction of expression was undertaken at different culture optical densities, and specific il-2 accumulation was found to decrease with increasing cell density at induction. induction at higher culture optical densities was also accompanied by decreased growth during induction and increased ace ...	1990	2180368
integration of bacteriophage lambda into the cryptic lambdoid prophages of escherichia coli.	bacteriophage lambda missing its chromosomal attachment site will integrate into reca+ escherichia coli k-12 and c at the sites of cryptic prophages. the specific regions in which these recombination events occur were identified in both lambda and the bacterial chromosomes. a noti restriction site on the prophage allowed its physical mapping. this allowed us to identify the locations of rac, qin, and qsr' cryptic prophages on the noti map of e. coli k-12 and, by analogy, to identify the cryptic ...	1990	2139644
vectors for constructing kan gene fusions: direct selection of mutations affecting is10 gene expression.	we describe several vectors for constructing translational fusions to the kan gene of tn5. fusions are constructed in vitro using multi-copy vectors containing unique cloning sites situated between upstream transcriptional terminators and a downstream kan gene lacking transcriptional and translational start signals. multi-copy fusions can be converted to single-copy chromosomal fusions by in vivo recombination with specific phage lambda vectors and vice versa. we find that kan fusions are often ...	1990	2165970
participation of rec genes of escherichia coli k 12 in w-reactivation of uv-irradiated phage lambda.	the effect of the recombinational deficiency on w-reactivation of uv-damaged phage lambda was explored. in this paper we show that w-reactivation is reduced by the recb21 and recf143 mutations after bleomycin (bm) and uv treatment. combination of these mutations in the recb21recf143 double mutant blocks w-reactivation completely after bm induction, but leaves residual w-reactivation ability after uv-irradiation, which is abolished by the introduction of uvrb deficiency (delta(uvrb-chla]. w-react ...	1990	1689458
temperature-inducible gene expression in bacillus subtilis mediated by the ci857-encoded repressor of bacteriophage lambda.	an efficient system to control the expression of cloned genes in bacillus subtilis was established by introducing the escherichia coli bacteriophage lambda ci857 repressor-pr promoter system into this host. a staphylokinase reporter gene (sak42d), which was fused to the lambda pr promoter was constitutively expressed in b. subtilis even when the ci857 gene was present on the same plasmid. s1 nuclease mapping of the transcription start point confirmed that the pr promoter was active in b. subtili ...	1990	1699846
celb, a gene coding for a bifunctional cellulase from the extreme thermophile "caldocellum saccharolyticum".	"caldocellum saccharolyticum" is an obligatory anaerobic thermophilic bacterium. a gene from this organism, designated celb, has been cloned in escherichia coli as part of a bacteriophage lambda gene library. this gene produces a thermostable cellulase that shows both endoglucanase and exoglucanase activities on test substrates and is able to degrade crystalline cellulose to glucose. the sequence of celb has homology with both exo- and endoglucanases described by others. it appears to have a cen ...	1990	2126700
site-directed insertion mutagenesis with cloned fragments in escherichia coli by p1 phage transduction.	a cloned gene with an insertion, which was made by introducing cat, was ligated to the cloning site of the phage lambda gt11. p1 phage grown on cells lysogenized with the recombinant lambda phage could transduce the mutant gene into the original site on the escherichia coli chromosome.	1990	2157955
a short dna sequence from lambda phage inhibits protein synthesis in escherichia coli rap.	the escherichia coli rap mutant inhibits vegetative growth of bacteriophage lambda. phage mutations termed bar, which overcome the rap defect, have been mapped to three genetic loci in the pl operon. plasmids with a lambda wild-type bar dna segment cloned downstream from an active promoter cannot be maintained in rap mutant bacteria. the viability of a rap mutant strain decreases rapidly after induction of transcription through bar regions present on plasmids. under these (restrictive) condition ...	1990	2147720
lysis protein s of phage lambda functions in saccharomyces cerevisiae.	the lambda s lysis gene was cloned into a saccharomyces cerevisiae expression vector under gal1 control. induction with galactose in s. cerevisiae terminated cell growth and prevented colony formation. several membrane proteins immunoreactive with anti-s antibody accumulated in the membranes, indicating that sodium dodecyl sulfate-resistant oligomers of s are formed, similar to those observed in the membranes of escherichia coli cells killed by expression of the s gene. these observations sugges ...	1990	2147680
stimulation of mutations suppressing the loss of replication control by small alcohols.	transient exposure of lysogenic escherichia coli cells to small alcohols stimulated the frequency of mutations suppressing the lethal loss of replication control from a prophage fragment of bacteriophage lambda. the stimulation in mutation frequency paralleled the effect of mutagenic agents, and in this sense the alcohols behaved as mutagens. 10-min treatments above distinct threshold concentrations at 23%, 18%, 10% and 4% (v/v) were required in order for methanol, ethanol, isopropanol and propa ...	1990	2143557
lambda vectors for stable cloned gene expression.	the bacteriophage lambda offers a unique opportunity concurrently to minimize segregational instability in recombinant systems by chromosomal integration of the cloned gene and to achieve high cloned gene expression during an abortive lytic phase. lysis leads approximately to a 100-fold amplification of the cloned gene. cell lysis in the lytic state is blocked by a specific mutation (sam), allowing the cell to maintain its integrity, and lambda dna packaging is blocked by other mutations (wam, e ...	1990	1368559
detection of protein-dna complex formation by time-resolved fluorescence depolarization of bound ethidium bromide.	we introduce the use of time-resolved fluorescence spectroscopy to probe the interaction between gene regulatory proteins and dna. changes in the decay kinetics of fluorescence polarization anisotropy of ethidium bromide bound to dna segments report changes in hydrodynamic volume and shape which occurs upon complex formation between protein and dna. we have used the decay of fluorescence polarization anisotropy as a spectroscopic handle on the interaction between several site-specific dna-bindin ...	1990	2291477
action of an rna site at a distance: role of the nut genetic signal in transcription antitermination by phage-lambda n gene product.	the n gene product of escherichia coli phage lambda is a transcriptional activator that captures the host rna polymerase and modifies it to a termination-resistant form, permitting gene expression in two large polycistronic operons of the phage genome. antitermination in vitro requires at least one host factor called nusa, which directly binds the n protein as well as rna polymerase, and also a transcribed cis-acting site known as nut, within which lies the hypothesized n-recognition signal, box ...	1990	2151659
the use of transgenic mice for short-term, in vivo mutagenicity testing.	in order to develop a short-term, in vivo assay to study the mutagenic effects of chemical exposure, transgenic mice were generated using a lambda shuttle vector containing a lacz target gene. following exposure to mutagens, this target can be rescued efficiently from genomic dna prepared from tissues of the treated mice using restriction minus, in vitro lambda phage packaging extract and restriction minus escherichia coli plating cultures. mutations in the target gene appear as colorless plaque ...	1990	2151115
characterization of the pilin gene of moraxella bovis dalton 2d and expression of pili from m. bovis in pseudomonas aeruginosa.	the pilin gene of moraxella bovis dalton 2d was isolated by cloning in pseudomonas aeruginosa. the nucleotide sequence of this gene encodes a prepilin of 156 amino acid residues. when high levels of pilin were expressed from the gene in p. aeruginosa, by using the pl promoter of bacteriophage lambda inserted upstream of the coding sequence, pili which were indistinguishable from pili of m. bovis were produced.	1990	1971258
a minor arginine trna mutant limits translation preferentially of a protein dependent on the cognate codon.	the escherichia coli argu gene encodes a rare arginine trna (anticodon ucu) that translates the similarly rare aga codon. the argu10(ts) mutation is a transition that changes the first nucleotide of the mature trna from g to a, presumably destabilizing the acceptor stem. this mutation, when present in haploid condition in the chromosome, reduces the growth rate at 30 degrees c and results in cessation of growth after 60 to 90 min at 43 degrees c. the mutation also preferentially limits (compared ...	1990	2139647
high level expression of genes cloned in phage lambda gt11.	plasmid cloning vectors have been constructed which allow genes originally cloned in lambda gt11 to be expressed at a high level in escherichia coli. they are based on the pembl and puc vectors, with the genes transcribed from the lac promoter. the ecori site in the vector has been altered to be in the same reading frame as the site used for cloning in lambda gt11. cloned proteins are expressed fused to a 2-kda leader sequence containing a run of six aparagine residues which considerably improve ...	1989	2527780
translesion synthesis is the main component of sos repair in bacteriophage lambda dna.	agents that interfere with dna replication in escherichia coli induce physiological adaptations that increase the probability of survival after dna damage and the frequency of mutants among the survivors (the sos response). such agents also increase the survival rate and mutation frequency of irradiated bacteriophage after infection of treated bacteria, a phenomenon known as weigle reactivation. in uv-irradiated single-stranded dna phage, weigle reactivation is thought to occur via induced, erro ...	1989	2527845
effects of all single base substitutions in the loop of boxb on antitermination of transcription by bacteriophage lambda's n protein.	the 'n' antitermination proteins of lambdoid bacteriophages are essential for overcoming multiple transcription terminators located within the major early operons of these phages (1). in order for n proteins to function, a genome sequence specifying n utilization, nut, must be located within an operon, between the promoter and the terminators (2). two components have been identified within nut: 8-base boxa, conserved among different phages and implicated in the recognition of host nusa protein, ...	1989	2527353
the escherichia coli protein, fis: specific binding to the ends of phage mu dna and modulation of phage growth.	we show, using gel retardation, that crude escherichia coli cell extracts contain a protein which binds specifically to dna fragments carrying either end of the phage mu genome. we have identified this protein as fis, a factor involved in several site-specific recombinational switches. furthermore, we show that induction of a mucts62 prophage in a fis lysogen occurs at a lower temperature than that of a wild-type strain, and that spontaneous induction of mucts62 is increased in the fis mutant. d ...	1989	2548061
phasing of protein-induced dna bends in a recombination complex.	many of the structures responsible for replication, transcription initiation and recombination arise from complex sets of protein-protein interactions and the folding of dna in three dimensions, with protein-induced bending of dna often playing an integral role. the magnitude and orientation of dna bending induced by various single proteins has been estimated by gel mobility shift methods and by modelling of crystallographic data. the site-specific recombination by which bacteriophage lambda (ph ...	1989	2528698
phage genetic sites involved in lambda growth inhibition by the escherichia coli rap mutant.	the rap mutation of escherichia coli prevents the growth of bacteriophage lambda. we have isolated phage mutants that compensate for the host deficiency. the mutations, named bar, were genetically located to three different loci of the lambda genome: bari in the attp site, barii in the ciii ea10 region, and bariii within or very near the imm434 region. the level of lambda leftward transcription correlates with rap exclusion. phage lambda mutants partially defective in the pl promoter or in pl-tr ...	1989	2523838
flanking dna-sequences contribute to the specific binding of ci-repressor and or1.	the binding of ci-repressor to a series of mutant operators containing or1 of the right operator of bacteriophage lambda was investigated. sites or2 and/or or3 were inactivated by either point or deletion mutations. the free energy of binding repressor to or1 in the wildtype operator, delta g1, is -13.7 +/- 0.3 kcal/mol. delta g1 determined for an or2- operator created by a single point mutation in or2 is -13.6 +/- 0.2 kcal/mol. in contrast, delta g1 for the binding of repressor to a cloned synt ...	1989	2525252
transcriptional activation of bacteriophage lambda dna replication in vitro: regulatory role of histone-like protein hu of escherichia coli.	initiation of bacteriophage lambda dna replication in vivo and in crude in vitro systems is strongly dependent on transcription at or near the lambda replication origin (ori lambda). through its capacity to prevent rna polymerase-mediated 'transcriptional activation' of lambda dna replication, the lambda ci repressor is capable of negatively regulating initiation of lambda dna replication, even when all required replication proteins are present. surprisingly, the strict requirement for transcrip ...	1989	2529119
cloning, sequencing and expression of the l-2-hydroxyisocaproate dehydrogenase-encoding gene of lactobacillus confusus in escherichia coli.	the gene (l-hicdh) encoding l-2-hydroxyisocaproate dehydrogenase (l-hicdh) from lactobacillus confusus was cloned in escherichia coli. a 69-mer oligodeoxyribonucleotide probe, derived to be complementary to the n-terminal amino acid (aa) coding sequence, was used for screening. the complete nucleotide (nt) sequence of the l-hicdh gene was determined. the 5'-end of the mrna was mapped by primer extension and the promoter identified. downstream from the l-hicdh gene is a typical rho-independent te ...	1989	2684788
the terminus of the escherichia coli chromosome is flanked by several polar replication pause sites.	replication of two small 'constrained' regions of the escherichia coli chromosome, one bordered by replication terminator t1 and the other by t2, displays normal velocity in the normal direction whereas it is much slower in the opposite direction (de massy et al., 1987). the presence of multiple polar terminators has been investigated, using a bacteriophage lambda derivative which provides a replication origin movable to predetermined loci and inducible on demand. the amount of dna made from thi ...	1989	2532703
bending of dna by gene-regulatory proteins: construction and use of a dna bending vector.	the binding of a protein to its specific sequence, borne on a dna fragment, retards the mobility of the fragment in a characteristic way during gel electrophoresis. if the protein induces bending in the dna, the contortion can also be monitored by gel electrophoresis, because the amount of retardation of the mobility of the dna-protein complex is dependent upon the position and the degree of the bend induced in the dna fragment [wu and crothers, nature 308 (1984) 509-513]. we have constructed a ...	1989	2533576
homologous recombination in prokaryotes: enzymes and controlling sites.	a common step in prokaryotic recombination appears to be the synapsis of the 3'-end of single-stranded dna with duplex dna to form a d-loop. the enzymatic mechanisms by which 3'-ends are produced and by which d-loops are converted into recombinant molecules are illustrated by proposed mechanisms of recombination by the escherichia coli recbcd pathway and the phage lambda red pathway. the enzymes promoting recombination and the special dna sites at which they act are emphasized. recombination by ...	1989	2534386
cloning and dna sequence of plasmid determinant iss, coding for increased serum survival and surface exclusion, which has homology with lambda dna.	escherichia coli k12 cells carrying a cloned 1.4 kb hindiii fragment from plasmid colv2-k94, showed increased survival in guinea pig serum. the recombinant plasmid also conferred group ii surface exclusion, i.e. the cells were reduced in recipient ability towards the incoming plasmid r538drd in conjugation experiments. southern blotting suggested homology with bacteriophage lambda dna and to the insertion element is2. determination of the dna sequence of the fragment demonstrated the presence of ...	1989	2546040
cysteine-22 and cysteine-38 are not essential for the functions of maltoporin (lamb protein).	maltoporin in the outer membrane of escherichia coli contains two cysteine residues, at positions 22 and 38 in the primary sequence. the role of these residues in determining structural stability, and their contributions to the maltoporin binding sites for maltodextrins and bacteriophage lambda, was investigated. site-directed mutagenesis was used to alter each of these residues to a serine. a double mutant lacking both cysteines was also isolated. none of the substitutions affected maltodextrin ...	1989	2693195
characterization of the cryptic lambdoid prophage dlp12 of escherichia coli and overlap of the dlp12 integrase gene with the trna gene argu.	the argu (dnay) gene of escherichia coli is located, in clockwise orientation, at 577.5 kilobases (kb) on the chromosome physical map. there was a cryptic prophage spanning the 2 kb immediately downstream of argu that consisted of sequences similar to the phage p22 int gene, a portion of the p22 xis gene, and portions of the exo, p, and ren genes of bacteriophage lambda. this cryptic prophage was designated dlp12, for defective lambdoid prophage at 12 min. immediately clockwise of dlp12 was the ...	1989	2553674
mapping of insertion elements is1, is2 and is3 on the escherichia coli k-12 chromosome. role of the insertion elements in formation of hfrs and f' factors and in rearrangement of bacterial chromosomes.	the chromosome of an escherichia coli k-12 strain w3110 contains seven copies of insertion element is1, 12 copies of is2 and six copies of is3. we determined the approximate locations of six copies of is1 (named is1a to is1f), ten copies of is2 (named is2a to is2j), and five copies of is3 (named is3a to is3e) on the w3110 chromosome by plaque hybridization using the "mini-set" of the lambda phage library that includes 476 clones carrying chromosomal segments that cover the w3110 chromosome almos ...	1989	2553981
location and molecular cloning of the structural gene for the deoxyguanosine triphosphate triphosphohydrolase of escherichia coli.	the structural gene for deoxyguanosine triphosphate triphosphohydrolase (dgtpase) (ec 3.1.5.1) and its regulator, opta, have been located on a lambda phage carrying a 17.5kb escherichia coli dna insert. the dna fragment has been excised and ligated into pbr325 and also transferred to another lambda vector. from the results of transduction and transformation experiments, we find that the structural gene for dgtpase is very closely linked to opta and dapd, which locates it at approximately 3.6 min ...	1989	2559296
overexpression of n antitermination proteins of bacteriophages lambda, 21, and p22: loss of n protein specificity.	the n protein of bacteriophage lambda (n lambda) modifies escherichia coli rna polymerase in such a way that it transcribes through termination signals, a process called antitermination. n antitermination normally occurs only if the template contains a specific utilization or nut site upstream of the terminators and only in the presence of host-encoded nus proteins. the lambda-related phages 21 and p22 produce n analogs, n21 and n22, but these require different nut sites and show a different pat ...	1989	2651405
bacteriophage t4 early promoter regions. consensus sequences of promoters and ribosome-binding sites.	twenty-nine early promoters from bacteriophage t4 and 14 early promoters from bacteriophage t6 were isolated using vector m13hdl17, a promoterless derivative of m13mp8 carrying a linker sequence, the bacteriophage lambda-terminator tr1, and the lacz' gene including part of its ribosome-binding site. the consensus sequence for the t4 promoters is: (sequence; see text). ribosome-binding sites of t4 share the sequence: 5'...g.ggaga..aa.atgaa.a...3' the consensus sequence of the t4 early promoter re ...	1989	2810355
generation of a large combinatorial library of the immunoglobulin repertoire in phage lambda.	a novel bacteriophage lambda vector system was used to express in escherichia coli a combinatorial library of fab fragments of the mouse antibody repertoire. the system allows rapid and easy identification of monoclonal fab fragments in a form suitable for genetic manipulation. it was possible to generate, in 2 weeks, large numbers of monoclonal fab fragments against a transition state analog hapten. the methods described may supersede present-day hybridoma technology and facilitate the producti ...	1989	2531466
characterization and vaccine potential of a novel recombinant coccidial antigen.	a cdna clone derived from sporulated oocysts of eimeria tenella and encoding the expression product gx3262 was identified using a monoclonal antibody raised against eimeria acervulina sporozoites. the cdna fragment containing the coccidial antigen gene was cloned in bacteriophage lambda gt11, transferred to a plasmid, and introduced into escherichia coli for analysis of the gene products. the strain carrying the plasmid produced gx3262 as part of a fusion protein consisting of the first 1,006 am ...	1989	2659532
genetic analysis of the lytic replicon of bacteriophage p1. ii. organization of replicon elements.	the region of bacteriophage p1 dna containing a lytic (vegetative) replicon has been identified by cloning p1 fragments into a phage lambda vector. we present the sequence of that replicon. using a novel fusion vector containing two p1 loxp recombination sites, we have developed a transformation assay for replicon function and have used that assay to identify some of the components of the p1 lytic replicon. among those components is a transcription promoter, p53, whose activity is essential for ...	1989	2661830
sumo15a: a lambda phasmid that permits easy selection for and against cloned inserts.	we report the construction of a phasmid vector, sumo15a, designed for recombination-based screening of recombinant dna libraries [seed, nucleic acids res. 11 (1983) 2427-2445]. this vector permits rapid selection in escherichia coli for homology-mediated integration and excision between homologous dna inserts cloned in a supf-carrying plasmid and in sumo15a. the region available for recombination spans the homologous sequence shared by the plasmid and the phasmid. supf is the selection tool that ...	1989	2559877
new derivatives of transposon tn5 suitable for mobilization of replicons, generation of operon fusions and induction of genes in gram-negative bacteria.	three types of new variants of the broad-host-range transposon tn5 are described. (i) tn5-mob derivatives with the new selective resistance (r) markers gmr, spr and tcr facilitate the efficient mobilization of replicons within a wide range of gram-negative bacteria. (ii) promoter probe transposons carry the promoterless reporter genes lacz, nptii, or luc, and nmr, gmr or tcr as selective markers. these transposons can be used to generate transcriptional fusions upon insertion, thus facilitating ...	1989	2551782
cloning and expression of the camp factor of group b streptococci in escherichia coli.	the genetic determinant of the camp factor from a strain of group b streptococcus (gbs; streptococcus agalactiae) was cloned in escherichia coli. total cell dna from the gbs strain r268 was used to construct a gene bank with bacteriophage lambda embl4 in the e. coli k-12 strain le392. recombinant phage plaques were detected by immunoblots by using a specific antiserum raised against purified camp factor. two hybrid phages showing expression of camp factor were identified. subcloning the camp gen ...	1988	3294187
aspects of the regulation of adenylate cyclase synthesis in escherichia coli k12.	in escherichia coli k12 expression of the adenylate cyclase gene is subject to multiple controls. in order to gain understanding of the regulation of adenylate cyclase synthesis, operon and protein fusions were constructed by in vitro recombination either into bacteriophage lambda or low-copy-number plasmids, or directly on the chromosome at the cya locus. the fusions were used in physiological experiments as probes to study transcriptional and translational controls of cya expression. it was fo ...	1988	3049935
mutations within the decoding site of escherichia coli 16s rrna: growth rate impairment, lethality and intragenic suppression.	several c----u transitions and small deletions were introduced into the conserved region centered on base c1400 in escherichia coli 16s rrna by in vitro mutagenesis. the mutations were placed within rrnb operons on multicopy plasmids under the transcriptional regulation of either the normal rrnb p1p2 promoters or the temperature-inducible pl promoter from bacteriophage lambda and introduced into e. coli hosts. when expressed from the p1p2 promoters, several of the mutant 16s rrnas impaired cell ...	1988	3047677