plasmid expression vector using the lambda late promoter.a plasmid expression vector called pqte1 based on the late promoter, pr', and positive control gene q of bacteriophage lambda has been constructed. this vector has unique cloning sites for placing exogenous dna under control of pr'. induction of expression of genes cloned into the pqte1 plasmid leads to massive overproduction of the gene products. also, transcription from the pr' promoter on pqte1 appears to be insensitive to polarity effects.19862940611
promoter recognition by escherichia coli rna polymerase. effects of substitutions in the spacer dna separating the -10 and -35 regions.a family of variants of the prm promoter of lambda phage was constructed, bearing nine base pair substitutions in a stretch of the spacer dna separating the contacted -10 and -35 regions. the substituted sequences were chosen for their potential to adopt structures different from those of average b-form dna and thus to affect the interaction of rna polymerase with the two contacted regions. characterization of the promoters in vitro and in vivo provides additional support for the lack of specifi ...19862942546
selective inhibition of escherichia coli recbc activities by plasmid-encoded gams function of phage lambda.the gam locus of bacteriophage lambda encompasses two coding sequences with the same reading frame and translational stop, one corresponding to an mr 11646 polypeptide (gams gene), the other to an mr 16349 polypeptide (gaml gene). a dna segment encoding gams but not gaml was placed under lambda pr promoter control (regulated by the cits857-coded repressor) on a multicopy plasmid, and an insertion mutation (gams201) was constructed. expression of gams+, but not gams201, inhibited escherichia coli ...19862943636
an improved method for the isolation of high yields of bacteriophage lambda dna. 19862947045
wavelength dependence for the induction of bacteriophage lambda by antitumor agent gilvocarcin v. 19862949330
chi-stimulated patches are heteroduplex, with recombinant information on the phage lambda r chain.generalized recombination in escherichia coli is elevated near chi sites. in vitro, recbcd enzyme can nick chi a few nucleotides 3' of the terminal gg of the chi sequence (5'-gctggtgg). the simplest model in which this nick at chi participates in chi function predicts that in phage lambda, chi-stimulated recombinants not crossed-over for flanking markers (patches) should be heteroduplex, with recombinant information on the lambda i chain. i report here that patches are heteroduplex, but that rec ...19872949853
structures of mismatched base pairs in dna and their recognition by the escherichia coli mismatch repair system.the escherichia coli mismatch repair system does not recognize and/or repair all mismatched base pairs with equal efficiency: whereas transition mismatches (g x t and a x c) are well repaired, the repair of some transversion mismatches (e.g. a x g or c x t) appears to depend on their position in heteroduplex dna of phage lambda. undecamers were synthesized and annealed to form heteroduplexes with a single base-pair mismatch in the centre and with the five base pairs flanking each side correspond ...19862951250
mutational analysis of integrase arm-type binding sites of bacteriophage lambda. integration and excision involve distinct interactions of integrase with arm-type sites.integrative recombination between specific attachment (att) regions of the bacteriophage lambda genome (attp) and the escherichia coli genome (attb) results in a prophage flanked by the hybrid recombinant sites attl and attr. each att site contains sequences to which proteins involved in recombination bind. using site-directed mutagenesis, we have constructed a related set of point mutations within each of the five int "arm-type" binding sites located within attp, attl and attr. footprint analys ...19862951525
large palindromes in the lambda phage genome are preserved in a rec+ host by inhibiting lambda dna replication.a large palindrome carried by phage lambda has been shown to prevent growth of the phage on a rec+ strain of escherichia coli. the phage do form plaques on recbc sbcb strains, but the palindrome is not stable--deletions that either destroy the palindrome or diminish its size overgrow the original engineered palindrome-containing phage. we have prepared stocks of lambda carrying a palindrome that is 2 x 4200 base pairs long. these phage stocks are produced by induction of a lysogen in which the t ...19872951734
mutations that improve the pre promoter of coliphage lambda.the dya5 mutation, a c----t change at position -43 of the lambda pre promoter, results in a twofold increase in pre activity in vivo. smaller increases in pre activity are found for the dya2 mutation, a t----c change at position -1 of pre, and the dya3 mutation, an a----g change at +5 of pre. the mutant pre promoters retain complete dependence on cii protein for activity. these observations argue, at least for pre-like promoters, that promoter activities are influenced by nucleotide sequences at ...19872953648
role of homology in site-specific recombination of bacteriophage lambda: evidence against joining of cohesive ends.bacteriophage lambda integration and excision take place at specific loci called attachment sites. earlier work has shown that efficient recombination requires the identical sequence to be present in both attachment sites throughout the seven-base-pair region between the points of strand exchange. a plausible model for the role of homology postulates that int, the site-specific recombinase, makes double-strand breaks at attachment sites such that each broken end has a short single-strand protrus ...19872954163
in vivo dna cloning with a mini-mu replicon cosmid and a helper lambda phage.a mini-mu bacteriophage, containing the cohesive-end packaging site (cos) from a lambda-phi 80 hybrid phage, a high-copy-number plasmid replicon, and a kanamycin-resistance gene for independent selection, was constructed to clone genes in vivo. this mini-mu element can be derepressed to transpose at a high frequency. dna segments that become flanked by copies of this mini-mu element in the same orientation can be packaged by a helper lambda phage. the resulting lambda lysate can be used to infec ...19872954879
construction of a dna-polymerase i overproducing plasmid and isolation of the enzyme.the pola gene of escherichia coli coding for dna polymerase i was cloned under the control of bacteriophage lambda promoter pl and gene n in a high copy number plasmid vector. the chromosomally located lambda cits repressor gene kept the synthesis of the pola gene product at 28 degrees c at a low level. raising the temperature to 43 degrees c resulted in inactivation of the repressor and overproduction of dna polymerase i, which could easily be purified to homogeneity.19872955618
mismatch repair of deaminated 5-methyl-cytosine.deamination of 5-methyl-cytosine in double-stranded dna produces a g-t mismatch. heteroduplexes of bacteriophage lambda dna containing a g-t mismatch at the site of a g-5-mec base-pair in one of the parental phages were constructed and used to transfect escherichia coli cells. genetic analysis of the progeny phages derived from such heteroduplexes suggests that, in e. coli, mismatches resulting from the deamination of 5-methyl-cytosine are repaired by a system requiring the e. coli dcm methylase ...19872956428
[reduced restriction of phage lambda in escherichia coli k-12 cells after gamma-irradiation].in gamma-irradiated cells of escherichia coli k-12 restriction alleviation of an unmodified phage lambda is only observed in ab1157 strain. no restriction alleviation by gamma-rays is registered in ab1157 mutants (rec a and ssb-1).19872957724
new mutations in the prm promoter of bacteriophage lambda.a prm-ci-lacz fusion inserted into the b2 region of bacteriophage lambda imm21 was used to isolate mutations in the lambda prm promoter. among the mutations causing defects in synthesis of both repressor (ci gene product) and beta-galactosidase, new promoter mutations were identified at positions -11 and -32 relative to the ci transcription start point. both mutations are changes in conserved (consensus) nucleotides in prm, but the mutation at -11, which alters a more highly conserved nucleotide ...19872958391
deletion analysis of the dna sequence required for the in vitro initiation of replication of bacteriophage lambda.supercoiled dna containing the replication origin of bacteriophage lambda can be replicated in vitro. this reaction requires purified lambda o and p replication proteins and a partially purified mixture of escherichia coli proteins (tsurimoto, t., and matsubara, k. (1982) proc. natl. acad. sci. u.s.a. 79, 7639-7643; wold, m. s., mallory, j.b., roberts, j. d., lebowitz, j. h., and mcmacken, r. (1982) proc. natl. acad. sci. u.s.a. 79, 6176-6180). the lambda origin region has four repeats of a 19-b ...19872958451
sequence of amino acids in lamb responsible for spontaneous ejection of bacteriophage lambda dna.the dna sequence of the promoter-distal half of lamb from shigella sonnei 3070 has been determined and compared with the known sequence for the escherichia coli k12 gene. the only predicted amino acid changes in this region of lamb, the receptor protein for bacteriophage lambda, lie between positions 381 and 390, where seven of the ten amino acids are altered. evidence is presented that indicates that this region is responsible for the ability of the s. sonnei receptor, but not the e. coli recep ...19872958635
isolation of escherichia coli rpob mutants resistant to killing by lambda cii protein and altered in pyre gene attenuation.escherichia coli mutants simultaneously resistant to rifampin and to the lethal effects of bacteriophage lambda cii protein were isolated. the sck mutant strains carry alterations in rpob that allow them to survive cii killing (thus the name sck), but that do not impair either the expression of cii or the activation by cii of the lambda promoters pe and pi. the sck-1, sck-2, and sck-3 mutations modify transcription termination. the growth of lambda, but not of the n-independent lambda variant, l ...19872959654
inhibition of streptomycin-dependent mutation in escherichia coli on the lytic growth of bacteriophage lambda.spontaneous streptomycin-dependent mutants (strda) were isolated from escherichia coli c600. on c600 strd, the lytic growth of phage lambda nam and lambda ci857 was inhibited. after e. coli lysogenic strain 1.1485 (lambda ci857) mutated to strda, induction of lambda was decreased greatly. on strda of e. coli strains c600 and 1.1485 (lambda ci857), the plating efficiencies and burst sizes of phage t4 and t7 remained normal. since strda is a mutation in the structural gene for ribosomal protein s1 ...19872960017
control of cloned gene expression by promoter inversion in vivo: construction of improved vectors with a multiple cloning site and the ptac promoter.we have constructed three gene-expression plasmids which contain (an) invertible promoter(s) and a multiple cloning site. we used either the plac promoter or the ptac-plac tandem promoters, the latter directing a more than fourfold increase in expression of the galk reporter gene in escherichia coli host. all these plasmids were derived from the pnh7a expression plasmid of podhajska et al. [gene 40 (1985) 163-168]. like pnh7a, these vectors have three novel properties: (i) in the 'off phase', th ...19872960590
molecular cloning of treponema pallidum outer envelope fibronectin binding proteins, p1 and p2.phages directing the synthesis of treponema pallidum fibronectin binding adhesin proteins, p1 and p2, were isolated from an embl-3 bacteriophage lambda library of t pallidum deoxyribonucleic acid (dna). the recombinant phages were identified using antisera generated to treponemal proteins purified in fibronectin-sepharose. recombinant p1 and p2 proteins possessed the same relative molecular weights as the native surface polypeptides of spirochaetes. the structural genes for these proteins were s ...19872962928
[phenomenon of restriction alleviation in escherichia coli strains].the alleviation of foreign dna restriction after treatment of cells by uv and gamma-rays or ocr+-gene product of t3, t7 phages has been studied. both uv and gamma-radiation were shown to induce in restriction alleviation of unmodified phage. the results of restriction alleviation caused by t3 and t7 ocr+-gene products have been evaluated by f-lac+ transfer efficiencies in heterologic crosses and plating of unmodified phage lambda. the phenomenon of restriction alleviation was established to depe ...19872963212
changes in dna base sequence induced by gamma-ray mutagenesis of lambda phage and prophage.mutations in the ci (repressor) gene were induced by gamma-ray irradiation of lambda phage and of prophage, and 121 mutations were sequenced. two-thirds of the mutations in irradiated phage assayed in reca host cells (no induction of the sos response) were g:c to a:t transitions; it is hypothesized that these may arise during dna replication from adenine mispairing with a cytosine product deaminated by irradiation. for irradiated phage assayed in host cells in which the sos response had been ind ...19882966755
role of the escherichia coli dnak and dnaj heat shock proteins in the initiation of bacteriophage lambda dna replication.we examined the role of two escherichia coli heat shock proteins, the dnak and dnaj gene products, during the initiation of lambda dv dna replication in vitro. using 14c-labeled lambda p protein we showed that the dnak and dnaj heat shock proteins function together to release lambda p protein from the preprimosomal complex consisting of lambda origin of replication-lambda o-lambda p-dnab protein. hydrolysis of atp, catalyzed presumably by dnak, is required during this reaction. substitution of d ...19882970643
cleavage of the cii protein of phage lambda by purified hfla protease: control of the switch between lysis and lysogeny.the activity of the cii protein of phage lambda is probably the critical controlling factor in the choice of the lytic or lysogenic pathway by an infecting virus. previous work has established that cii activity is regulated through the turnover of cii protein; the products of the hfla and hflb loci of escherichia coli are needed for a degradative reaction, and lambda ciii functions in stabilizing cii. by using the cloned hfla locus, we have purified a cii-cleaving enzyme that we term hfla. purif ...19882973057
rapid isolation of lambda phage dna in micro- and macro-variants. 19882974539
coliphage lambda to terminator lowers the stability of messenger rna in escherichia coli hosts.the effects of the transcription terminators to and tfd on the overall high-level expression of a human interferon-beta gene (ifn-beta) in escherichia coli hosts were compared. deletion mapping shows that mrna lability is caused by sequences at or near the lambda terminator to stem-loop structure. extensive rna secondary structure in this region indicates a potential rnase iii cleavage/binding site. in rnase iii- e. coli hosts, ifn-beta synthesis is indeed considerably enhanced. the bacteriophag ...19882977353
evidence that the outer membrane protein gene nmpc of escherichia coli k-12 lies within the defective qsr' prophage.recombinants between phage lambda and the defective qsr' prophage of escherichia coli k-12 were made in an nmpc (p+) mutant strain and in the nmpc+ parent. the outer membrane of strains lysogenic for recombinant qsr' phage derived from the nmpc (p+) strain contained a new protein identical in electrophoretic mobility to the nmpc porin and to the lc porin encoded by phage pa-2. lysogens of qsr' recombinants from the nmpc+ strain and lysogens of lambda p4, which carries the qsr' region, did not pr ...19852984173
the phi 80 and p22 attachment sites. primary structure and interaction with escherichia coli integration host factor.although the lambdoid bacteriophage phi 80 and p22 possess site-specific recombination systems analogous to bacteriophage lambda, they have different attachment (att) site specificities. we have identified and determined the nucleotide sequences of the att sites of phi 80 and p22 and have examined the interaction of these sites with purified escherichia coli integration host factor (ihf). the sizes of the homologous core regions of the att sites vary greatly: p22 has a 46-base pair core, while p ...19852984205
phasmid vectors for identification of genes by complementation of escherichia coli mutants.a bacteriophage lambda cloning vector was designed to facilitate the isolation of genes from procaryotic organisms by complementation of escherichia coli mutants. this vector, lambda se4, was constructed by attaching a very-low-copy-number replication system (from the plasmid nr1) and a spectinomycin resistance gene to the left arm of lambda 1059 (karn et al., proc. natl. acad. sci. u.s.a. 77:5172-5176, 1980). this phasmid cloning vector is capable of growing lytically as a phage in a nonimmune ...19852985547
the dna restriction endonuclease of escherichia coli b. ii. further studies of the structure of dna intermediates and products.the dna intermediates and final products formed by the type i restriction endonuclease, ecob, were further characterized. dna cleaved on only one strand (hemi-restricted dna) contains gaps of approximately 70-100 nucleotides, while the fully restricted products contain 3'-single-stranded tails averaging approximately 70-100 nucleotides for each strand cleaved. the gaps and tails are formed with the release of an equal number of nucleotides as small oligonucleotides that are soluble in acid. afte ...19852985610
plasmid vectors designed for the analysis of transcription termination signals.we have constructed synthetic operons in which two genes (cat and lacz or cat and galk) were placed in tandem under the control of the bacteriophage lambda olpl operator and promoter. restriction sites were introduced between the promoter and the proximal cat gene or between the cat and lacz or galk genes. in the latter case, introduction of a transcriptional terminator between the two structural genes should affect only the distal gene. thus, following induction, the expression of the cat gene ...19852998929
f'-coded, temperature-sensitive lambda ci857 repressor gene for easy construction and regulation of lambda promoter-dependent expression systems.we describe the construction and properties of an f' factor which carries the temperature-sensitive ci857 allele of the repressor gene of coliphage lambda and which lacks the lambda cro function. this episome can easily be transferred to any f- and f' escherichia coli strain, thus facilitating the construction and regulation of lambda promoter-dependent expression systems without the use of defective prophages.19852999084
site-directed cleavage of immunoglobulin gene segments by lymphoid cell study the enzyme(s) involved in the site-specific recombination of immunoglobulin (ig) gene segments, we designed an assay to detect v-j joining in vitro. the dna from a hybrid phage (lambda vjck) containing the vk41 gene segment separated by a 6-kilobase spacer region from the entire j-ck sequence was incubated with lymphoid cell extracts and packaged in vitro. phages carrying genomic deletions were selected by screening for ethylenediaminetetraacetic acid resistance. although no site-specif ...19853000549
[deletion of late genes of the prophage lambda].the deletions in tandem prophage lambda appear with high frequency (to 10%) in rec a- strain of escherichia coli. the deletions were shown by marker rescue and hybridization of fragments of dna on nitrocellulose filters with nick-translated phage lambda dna localized only in prophage area. right and left att sites are not involved. the majority of defective lysogens had all regulatory regions and deletions of late structural genes. these strains may be used for construction of the host-vector sy ...19853001508
the nucleotide sequence of the escherichia coli k12 dnaj+ gene. a gene that encodes a heat shock protein.the escherichia coli dnaj gene product is required for bacteriophage lambda dna replication at all temperatures. it is also essential for bacterial viability in at least some conditions, since mutations in it result in temperature-sensitive bacterial growth. we have previously cloned the dnaj gene and shown that its product migrates as a mr 37,000 polypeptide under denaturing conditions. here we present the primary dna sequence of the dnaj gene. it codes for a processed basic protein (63 basic a ...19863003085
plasmids supplying the q-qut-controlled gam function permit growth of lambda red- gam- (fec-) bacteriophages on reca- hosts.a plasmid, pgam, has been constructed which expresses the phage lambda gene, gam, under the control of the lambda late promoter, p'r, contained in a form of a p'r-qut-t'r1 module. lambda red- gam-, which normally do not grow on reca- hosts, are able to grow on reca- hosts containing pgam, because their q function can turn on the gam gene expression. this facilitates cloning with lambda red- gam- vectors in reca- hosts.19853005123
genetic selection of lambda phage clones from a gene clonotheque.a genetic procedure for selection of specific lambda clones, by homologous recombination between lambda clones from a gene clonotheque and sequences cloned into a plasmid, was developed. resulting clones are isolated in transduction experiments by plating infected escherichia coli cells under conditions selecting for the antibiotic resistance marker carried by the plasmid. the feasibility of the method was demonstrated in a model test system as well as by isolation of alpha-interferon-specific s ...19863016484
cloning of micrococcus luteus 3-methyladenine-dna glycosylase genes in escherichia coli.the 3-methyladenine-dna glycosylase (m3adg) excises 3-methyladenine (m3a) residues formed in dna after treatment with alkylating agents. in escherichia coli, the repair of this type of damage depends on the products of the genes taga and/or alka, which code for m3adg i (20 kda) and ii (30 kda), respectively. the taga- and alka--single mutants are sensitive to alkylating agents, the double mutant much more so. we have cloned two genes of micrococcus luteus that can partly substitute the function ...19863019831
the fos gene product undergoes extensive post-translational modification in eukaryotic but not in prokaryotic investigate the properties of the fos oncogene, we have constructed bacterial and yeast vectors which express the entire fos-coded protein (fos) and two c-terminal deletion products. in escherichia coli, fos proteins were expressed from the phage lambda pl promotor under the control of the temperature-sensitive lambda repressor. in vitro transcription/translation studies indicate that these vectors produce fos proteins of the expected sizes. however, in vivo, fos protein accumulation is obser ...19863019839
[plasmid vector for gene expression containing the temperature-dependent promoter of phage lambda p'r dna]. 19863026761
the production of generalized transducing phage by bacteriophage lambda.generalized transduction has for about 30 years been a major tool in the genetic manipulation of bacterial chromosomes. however, throughout that time little progress has been made in understanding how generalized transducing particles are produced. the experiments presented in this paper use phage lambda to assess some of the factors that affect that process. the results of those experiments indicate: the production of generalized transducing particles by bacteriophage lambda is inhibited by the ...19863034738
regulation of dnab function in dna replication in escherichia coli by dnac and lambda p gene products.the dnab protein of escherichia coli, a multifunctional dna-dependent ribonucleotide triphosphatase and datpase, cross-links to atp on ultraviolet irradiation under conditions that support rntpase and datpase activities of dnab protein. the covalent cross-linking to atp is specifically inhibited by ribonucleotides and datp. tryptic peptide mapping demonstrates that atp cross-links to only the 33-kda tryptic fragment (fragment ii) of dnab protein. the presence of single-stranded dna alters the co ...19873034907
sequence determination and comparison of the exfoliative toxin a and toxin b genes from staphylococcus aureus.the dna encoding the exfoliative toxin a gene (eta) of staphylococcus aureus was cloned into bacteriophage lambda gt11 and subsequently into plasmid pli50 on a 1,391-base-pair dna fragment of the chromosome. exfoliative toxin a is expressed in the escherichia coli genetic background, is similar in length to the toxin purified from culture medium, and is biologically active in an animal assay. the nucleotide sequence of the dna fragment containing the gene was determined. the protein deduced from ...19873040666
identification of polypeptides encoded by an escherichia coli locus (hfla) that governs the lysis-lysogeny decision of bacteriophage lambda.we report the cloning of the escherichia coli hfla locus, which governs stability of phage lambda cii protein and which has been proposed to encode or regulate a cii-specific protease. the hfla locus was cloned on an 18-kilobase dna fragment by selecting for plasmids that carry the neighboring pura gene. the boundaries of hfla were delimited by analysis of deletions and insertions constructed in vitro and by use of transposon tn1000. maxicell analysis of the proteins encoded by the hfla-containi ...19873040675
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 ...19883047677
aspects of the regulation of adenylate cyclase synthesis in escherichia coli 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 ...19883049935
molecular cloning and genetic analysis of the determinant for gamma-lysin, a two-component toxin of staphylococcus aureus.the gamma-lysin determinant of staphylococcus aureus strain smith 5r has been cloned in phage lambda and plasmid vectors in escherichia coli. genetic evidence is presented which demonstrates that gamma-lysin requires the co-operative action of two polypeptides expressed by the closely linked hlga and hlgb genes. recombinants expressed haemolytic activity in agarose medium but not in agar, a known property of gamma-lysin. haemolysis was inhibited by antiserum raised against the 32 kda component o ...19883075655
regulation of the aroh operon of escherichia coli by the tryptophan repressor.regulation of expression of aroh, the structural gene for the tryptophan-sensitive 3-deoxy-d-arabinoheptulosonic acid-7-phosphate synthetase, by the tryptophan repressor and its corepressor, l-tryptophan, was studied in vivo by using aroh-lacz fusions. protein and operon fusions were constructed on multicopy plasmids and subsequently crossed in single copy to the bacterial chromosome via the specialized transducing bacteriophage lambda rz5. analysis of the resulting lysogens demonstrated that ar ...19873106331
effect of antiserum against aflatoxin b1-bovine serum albumin complex on aflatoxin b1-induced lysogenesis.escherichia coli k12 bacteria lysogenic for the lambda phage were used to study the effect of antiserum against aflatoxin b1-induced lysogenesis. the antiserum was obtained from rabbits immunized with water in oil emulsion of aflatoxin b1-bovine serum albumin complex (afb1-bsa). a marked reduction in the degree of lysogenesis was observed when the antiserum was added to the reaction medium prior to microsomal enzyme activation of aflatoxin b1. there was no detectable effect when the antiserum wa ...19883140005
mutant trp repressors with new dna-binding specificities.oligonucleotide-directed mutagenesis of the codons for glutamine-68 (gln68), lysine-72 (lys72), isoleucine-79 (ile79), alanine-80 (ala80), and threonine-81 (thr81) of the escherichia coli trpr (tryptophan aporepressor) gene was used to make mutant repressors with each of 36 different amino acid changes. mutant repressors were tested for binding to each member of a set of 28 different operators closely related to the consensus trp operator. of the 36 mutant repressors, 11 bind a subset of the 28 ...19883140377
salmonella typhimurium lt2 metf operator mutations.using an escherichia coli lac deletion strain lysogenized with lambda phage carrying a metf-lacz gene fusion (lambda flac), in which beta-galactosidase levels are dependent on metf gene expression, cis-acting mutations were isolated that affect regulation of the salmonella typhimurium metf gene. the mutations were located in a region previously defined as the metf operator by its similarity to the e. coli metf operator sequence. regulation of the metf gene was examined by measuring beta-galactos ...19883147373
a phi 80 function inhibitory for growth of lambdoid phage in him mutants of escherichia coli deficient in integration host factor. ii. physiological analysis of the abortive infection.derivatives of phage lambda with the rightmost 3% of the genome (the qsr region) from the related phage phi 80 fail to grow at low temperatures (e.g., 32 degrees) in escherichia coli hosts deficient in either protein component of ihf (integration host factor), the products of the hima and hip/himd genes. the abortive infection of lambda (qsr)80 in mutants defective for ihf was studied in detail. this infection is characterized by a lack of cell lysis and an inhibition of phage dna replication af ...19853155886
involvement of the htpr gene product of escherichia coli in phage lambda development.growth of phage lambda at high temperature requires a functional htpr host gene. the stages of the phage growth cycle shown to be dependent on htpr gene function include prophage excision and particle morphogenesis. two types of morphogenetic abnormalities have been detected. one is a defect in phage tail assembly that results from a deficiency in tail fibers even though gpj is produced. the severity of this defect is phage-strain specific. the second morphogenetic defect is less clearly defined ...19853156447
efficient modification of e. coli rna polymerase in vitro by the n gene transcription antitermination protein of bacteriophage lambda.the n gene protein of bacteriophage lambda prevents termination of transcription by e. coli rna polymerase. we describe here the conditions of a cell-free reaction system in which pure n stimulates net transcription up to tenfold and therefore nearly stoichiometrically modifies transcribing rna polymerase molecules. the reaction contains micrococcal nuclease-treated s100 extract derived from e. coli and a plasmid template dna containing the lambda early promoter pl, the n utilization site nutl, ...19853158883
direct selection of mutations reducing transcription or translation of the reca gene of escherichia coli with a reca-lacz protein fusion.when a reca-lacz protein fusion was cloned into phage lambda, the resulting transducing phage grew normally on wild-type escherichia coli, but its growth was severely inhibited in lexa(def) mutant strains that express reca constitutively at high levels. mutants of the transducing phage that grew on the lexa(def) strains were isolated and were found to affect production of the reca-beta-galactosidase hybrid protein. most mutants, including a number of nonsense mutants, were phenotypically lacz-. ...19853160689
gene fusions to the ptsm/pel locus of escherichia coli.we have constructed gene fusions between ptsm/pel and lacz. these fusions affect both phenotypes assigned to the ptsm/pel locus (at 40 min), namely, no growth on mannose or glucosamine and inhibition of the penetration of bacteriophage lambda dna, as well as that of other lambdoid phages such as hy-2. since the lacz gene fusions are insertion mutations that abolish target gene function by disrupting the linear contiguity of the gene, it would appear that ptsm and pel are either the same gene or ...19853162078
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 ...19883294187
n-methyl-n'-nitro-n-nitrosoguanidine-induced resistance to ionizing radiation.n-methyl-n'-nitro-n-nitrosoguanidine (mnng) pretreatments increase the resistance of escherichia coli to gamma-radiation. the increased resistance is dependent on functional pola, reca, recb, recc, and lexa genes and is partly dependent on recn. the mnng-induced resistance is additive to resistance induced by pretreatment with gamma-radiation but not by increases induced by hydrogen peroxide. the mnng-induced resistance occurs in adaptive response mutants and at pretreatment levels of mnng that ...19873295481
cloning structural genes for treponema pallidum immunogens and characterisation of recombinant treponemal surface protein, p2 (p2 star).a genomic library consisting of partially digested 10 to 20 kilobase pair fragments of treponema pallidum deoxyribonucleic acid (dna) was constructed using bacteriophage lambda embl-3 as the vector. positive clones expressing t pallidum antigens were detected with sera from experimentally infected rabbits. treponemal proteins ranging in molecular weight from 37,000 daltons to 120,000 daltons were identified by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and immunoblotting of phage ...19873315959
extracellular production of cloned alpha-amylase by escherichia coli.overexpression of bacillus stearothermophilus gene coding for thermostable alpha-amylase in escherichia coli was shown to cause outer-membrane damage leading to extracellular location of periplasmic proteins. prolonged high expression of the alpha-amylase gene under laczpo control eventually also lysed cells. surprisingly, expression controlled by the pl promoter of phage lambda allowed specific release of periplasmic proteins into the growth medium without total cell lysis. accumulation of alph ...19873327755
cloning and purification of a unique lysozyme produced by bacillus phage phi 29.a dna fragment of the bacteriophage phi 29 chromosome, encoding the entire sequence of phi 29 gene 15, has been cloned into the escherichia coli expression vector pplc245 under the control of the phage lambda major leftward promoter, pl. upon heat induction, a protein with an apparent molecular mass of 26 kda was overproduced. the molecular mass of this protein corresponds to the 28 kda predicted for the product of gene 15 from its nucleotide sequence. the overproduced protein has been purified ...19873469652
cloning and expression of bradyrhizobium japonicum uptake hydrogenase structural genes in escherichia identify the structural genes for the components of bradyrhizobium japonicum uptake hydrogenase (mr 60,000 and 30,000), we have expressed these genes in escherichia coli and shown that the products cross-react with antibodies to the respective hydrogenase subunits. we constructed subclones of overlapping dna fragments from an uptake hydrogenase-complementing cosmid, phu52 [lambert, g. r., cantrell, m. a., hanus, f. j., russell, s. a., haddad, k. r. & evans, h. j. (1985) proc. natl. acad. sci. ...19863532119
enhancement of transcriptional activity of the escherichia coli trp promoter by upstream a + t-rich regions.the escherichia coli trp promoter has two a + t-rich blocks in the upstream region. the deletion of the segments containing these blocks resulted in a decrease in promoter strength. by replacing the upstream region of trp promoter with one or two large a + t-rich blocks of the major leftward lambda promoter (pl), modified trp promoters (designated let) were constructed, and the transcriptional activities of these promoters towards the expression of the human interferon-gamma gene were measured. ...19863533725
construction of plasmid vectors for the detection of streptococcal promoters.plasmid vectors have been constructed for detecting dna fragments that exhibit promoter activity in streptococcus sanguis. the plasmids are able to replicate in both s. sanguis and escherichia coli, and contain an erythromycin resistance marker which is expressed in both hosts. selection for promoter activity is dependent upon the insertion of appropriate dna fragments upstream from a promoterless chloramphenicol acetyl transferase gene (cat) from staphylococcus aureus. to facilitate this insert ...19863536665
expression of the bacteriophage t4 denv structural gene in escherichia coli.the expression of the t4 denv gene, which previously had been cloned in plasmid constructs downstream of the bacteriophage lambda hybrid promoter-operator olpr, was analyzed under a variety of growth parameters. expression of the denv gene product, endonuclease v, was confirmed in dna repair-deficient escherichia coli (uvra reca) by western blot analyses and by enhancements of resistance to uv irradiation.19863536845
repressor for the sn-glycerol-3-phosphate regulon of escherichia coli k-12: cloning of the glpr gene and identification of its product.the glpr gene encoding the repressor for the glp regulon of escherichia coli was cloned from a library of hindiii dna fragments established in bacteriophage lambda. phages harboring glpr were isolated by selection for sn-glycerol-3-phosphate dehydrogenase function encoded by glpd, which is adjacent to glpr on the e. coli linkage map. restriction endonuclease analysis and recloning of dna fragments localized glpr to a 3-kilobase-pair ecori-sali segment of dna. strains exhibiting constitutive expr ...19853881401
cloning and genetic analysis of serotype 5 m protein determinant of group a streptococci: evidence for multiple copies of the m5 determinant in the streptococcus pyogenes genome.a gene bank of group a streptococcus strain manfredo (m protein serotype 5) was constructed with a bacteriophage lambda vector-escherichia coli k-12 host system and screened by immunoblotting hybrid phage plaques with antisera raised to purified pep m5 (serotype 5 m protein fragment released from the streptococcal cell surface by pepsin). hybrid phage expressing m5 antigen (lambda m5) were detected in the gene bank at an unexpectedly high frequency. the cloned streptococcal dna sequences from on ...19853884510
overproduction of staphylokinase in escherichia coli and its characterization.a recombinant plasmid which directs the overproduction in escherichia coli of staphylokinase from staphylococcus aureus has been constructed by placing the staphylokinase gene, sak, under the control of bacteriophage lambda pr promoter in the plasmid. when an e. coli strain having the plasmid was induced, the staphylokinase activity in the periplasmic fraction increased about 60-fold and the 15.5-kda protein corresponding to the mature form reached about 25% of the periplasmic proteins. at the s ...19853891339
an escherichia coli mutant which inhibits the injection of phage lambda dna. 19744132241
isolation of the bacteriophage lambda receptor from escherichia coli.a factor which inactivates the phage lambda can be extracted from escherichia coli. this factor is a protein and is located in the outer membrane of the bacterial envelope. it is found in extracts of strains which are sensitive to phage lambda, but not in extracts of strains specifically resistant to this phage. we conclude that this factor is the lambda receptor, responsible for the specific adsorption of the phage lambda to e. coli cells. a partial purification of the lambda receptor is descri ...19734201774
purification and base composition analysis of phage lambda early promoters.rna-polymerase of escherichia coli was allowed to bind to dna of phage lambda in the absence of precursors. the resulting complex was excised by nuclease digestion and the protected dna was recovered by phenol-extraction and ethanol precipitation. acrylamide gel electrophoresis of protected dna fragments reveals the existence of two distinct oligonucleotide peaks corresponding, respectively, to 45-52 and 7-10 nucleotide residues along with species of intermediate sizes. peak i molecules have two ...19714332016
synthesis of bacteriophage lambda proteins in vitro. 19724343946
lambda phage transcription in human fibroblasts. 19724551993
on some genetic aspects of phage lambda resistance in e. coli k12.most mutations rendering e. coli k12 resistant to phage lambda, map in two genetic regions mala and malb.-the malb region contains a gene lamb specifically involved in the lambda receptor synthesis. twenty-one independent lamb mutations studied by complementation belonged to a single cistron. this makes it very likely that lamb is monocistronic. among the lamb mutants some are still sensitive to a host range mutant of phage lambda. mutations mapping in a proximal gene essential for maltose metab ...19724558324
interaction of the recombination pathways of bacteriophage lambda and host escherichia coli: effects on lambda recombination. 19724563260
methylation pattern of lambda deoxyribonucleic acid.deoxyribonucleic acid (dna) extracted from phage lambda grown on escherichia coli k-12 strain w4032 had 113 +/- 10 5-methylcytosine residues and 215 +/- 20 6-methyl adenine residues per genome, as determined by three independent methods. these methylated nucleotides were distributed equally among the two strands of lambda dna. shearing of double-stranded dna to half-length fragments revealed a slight deficiency of 5-methyl cytosine in the 55% guanine plus cytosine half. shearing the dna to fragm ...19724564587
properties of a mutant of escherichia coli defective in bacteriophage lambda head formation (groe). i. initial characterization. 19734578098
a new effect of the rex gene of phage lambda: premature lysis after infection by phage t1. 19734583305
prophage lambda at unusual chromosomal locations. ii. mutations induced by bacteriophage lambda in escherichia coli k12. 19734587404
radiochemical purification of bacteriophage lambda integrase. 19744589454
genetic analysis of bacteriophage lambda strains which transduce the genes for leucine biosynthesis. 19734591366
control of gene expression in bacteriophage lambda. 19734593306
preparation of plasmid lambda dv from bacteriophage lambda: role of promoter-operator in the plasmid replicon.a technique has been described for selection of bacteria carrying plasmid lambdadv. with this technique, the effects of mutations in the promoter-operators were compared on the production and perpetuation of the plasmid. it was found that "left" promoter-operator that controls leftward gene expressions can be deleted from the plasmid genome. some mutations of "right" promoter-operator (pror) that controls expression of genes tof, o, and p affect the stability of the plasmid. however, the plasmid ...19744595898
bacteriophage lambda derivatives carrying two copies of the cohesive end site. 19744598361
n protein causes the lambda dv plasmid to inhibit heteroimmune phage lambda imm434 growth and stimulates lambda dv replication. 19744608877
interaction of escherichia coli o55 hybrids with bacteriophage lambda. a new type of host specificity between escherichia coli o55 and urinary escherichia coli. 19744609858
new method for large-scale preparation of covalently closed lambda dna molecules.a combination of mutations in bacteriophage lambda and its host escherichia coli k-12 provides a convenient system for the isolation of large quantities of covalently closed circular dna molecules. we describe two procedures for the large scale preparation of lambda dna in the duplex circular form.19744610176
specific hydrolysis of the cohesive ends of bacteriophage lambda-deoxyribonucleic acid by micrococcus luteus ultraviolet exonuclease. 19744612037
inactivation of transfecting dna by physical and chemical agents: influence of genotypes of phage lambda and host cells. 19744612352
characterization of bacteriophage lambda reverse as an escherichia coli phage carrying a unique set of host-derived recombination functions. 19744616090
bacteriophage lambda fii gene protein: role in head assembly. 19744616093
repair mechanisms involved in the recovery of phage lambda from ultraviolet damage. 19734618990
[effect of mutations in genes n and p on the ability of uv-irradiated phage lambda to induce synthesis of colicin e1 in escherichia coli]. 19744830043
[the effect of cystamine and magnesium ions on the induction of moderate lambda phage by x-rays]. 19684869266
[comparative characteristics of oxygen consumption and electrokinetic potential of escherichia coli under the effect of moderate and virulent coliphage. 1. effect of the induction of lambda-phage production on oxygen consumption and electrophoretic motility of e. coli k12 lambda]. 19684873878
studies on radiation-sensitive mutants of e. coli. ii. breakage and repair of ultraviolet irradiated intracellular dna of phage lambda. 19684879098
antigenic differences between wild type and mutants of bacteriophage lambda. 19694887235
the lysozyme of bacteriophage lambda. 3. ordering the cyanogen bromide peptides. 19694889462
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