regulation of glutamine synthetase formation in escherichia coli: characterization of mutants lacking the uridylyltransferase.a lambda phage (lambdank55) carrying the translocatable element tn10, conferring tetracycline resistance (tetr), has been utilized to isolate glutamine auxotrophs of escherichia coli k-12. such strains lack uridylyltransferase as a result of an insertion of the tn10 element in the glnd gene. the glnd::tn10 insertion has been mapped at min 4 on the e. coli chromosome and 98% contransducible by phage p1 with dapd. a lambda transducing phage carrying the glnd gene has been identified. a glnd::tn10 ...197826660
the organization of ribosomal rna genes in the mitochondrial dna of tetrahymena pyriformis strain st.1. we have constructed a physical map of the mtdna of tetrahymena pyriformis strain st using the restriction endonucleases ecori, psti, saci, hindiii and hhai. 2. hybridization of mitochondrial 21 s and 14 s ribosomal rna to restriction fragments of strain st mtdna shows that this dna contains two 21-s and only one 14-s ribosomal rna genes. by s1 nuclease treatment of briefly renatured single-stranded dna the terminal duplication-inversion previously detected in this dna (arnberg et al. (1975) b ...1978102353
organization of the ribosomal ribonucleic acid gene cluster of neurospora crassa by means of restriction endonucleases and cloning in bacteriophage lambda [proceedings]. 1978154424
[cloning of the hepatitis b virus genome in escherichia coli].the whole genome of the hepatitis b virus (dane particles) was inserted in vitro in the genome of the bacteriophage lambda gtwes . lambda b. the recombinant dna molecule was cloned in e. coli. amplification of the hybrid bacteriophage enables the preparation of large amounts of hepatitis b virus dna. the possibilities offered by the utilization of this recombinant bacteriophage are discussed.1978158442
site-specific recombination of bacteriophage lambda: the role of host gene products. 1979158465
nucleotide sequence of a secondary attachment site for bacteriophage lambda on the escherichia coli chromosome.the nucleotide sequence of a secondary attachment site for bacteriophage lambda was determined in a region near the rrnb gene at 88 min on the e. coli chromosome. the sequence has a 8 base pair interrupted homology gct tttta to the common core of the primary attachment site (attb) and the corresponding phage sequence (attp). the site of crossover during integration lies probably between nucleotides -3 and +1. the flanking regions have no obvious homology to the arms of either attp or attb.1979160033
a rightward promoter to the left of the att site of lambda phage dna: possible participant in site-specific recombination.the binding has been studied of escherichia coli rna-polymerase to the fragments of lambda dna obtained by digestion with restriction endonucleases bsui, hindiii, bamhi, ecori and hindii + iii. there are at least six sites of rna-polymerase binding in the b2-region. in vitro transcription of those bsui-fragments of the b2-region which contain six binding sites is rightward. therefore, the fragments contain promoters rather than mere rna-polymerase binding sites. one of the promoters of the b2 re ...1979160359
the fate of phage lambda dna in lambda-infected minicells.the fate of phage lambda dna in lambda-infected escherichia coli minicells harboring the plasmid cole1, and in plasmid-free minicells, were studied. binding of lambda dna to the minicell membrane, and formation of the supercoiled covalently-closed circular structure has been demonstrated. phage infection abolishes plasmid dna synthesis. only a very slight, non-replicative lambda dna synthesis occurs, soon after infection. this synthesis is associated with fragments of lambda dna arising during, ...1979161175
multiple steps during the interaction between coliphage lambda and its receptor protein in vitro. 1976180667
molecular cloning of polyoma virus dna in escherichia coli: oncogenicity testing in hamsters.inoculation of suckling hamsters with 2 x 10(8) live cells of escherichia coli k12 strain chi1776, carrying the complete genome of polyoma virus in a recombinant plasmid, failed to induce tumors in any of 32 recipients. also, lambda phage dna and particles with a monomeric insert of polyoma dna did not induce tumors. purified recombinant plasmid dna, as well as phage particles and dna containing a head-to-tail dimer of polyoma dna, showed a low degree of oncogenicity, comparable to that of polyo ...1979224458
[integration of lambda phage (author's transl)]. 1979233426
identification of the n gene protein of bacteriophage lambda.the n gene protein, pn, of bacteriophage lambda stimulates early gene transcription by allowing mrna chain elongation to proceed into genes distal to transcription termination sites normally recognized by the escherichia coli transcription termination protein rho. pn has previously eluded detection on sodium dodecyl sulfate/polyacrylamide gels because of its small size, its instability, and the difficulty of distinguishing pn itself both from host proteins and from other early lambda proteins wh ...1978276863
enzymatic breakage of the cohesive end site of phage lambda dna: terminase (ter) in vitro system is described for measuring the endonucleolytic conversion of the phage lambda cohesive end sites in concatemeric dna to the cohesive chromosomal ends of the mature molecule. this enzymic process, known as the ter reaction, is catalyzed by purified lambda a gene protein. the reaction is markedly stimulated by atp, mg2+, spermidine, and one or more uncharacterized factors present in extracts of uninfected escherichia coli cells. in vitro, the ter reaction proceeds in the absence ...1978279909
a single base-pair change creates a chi recombinational hotspot in bacteriophage lambda.x4+ mutations, responsible for the chi phenotype in phage lambda, locally increase the rate of recombination promoted by the escherichia coli recombination system (rec). x+ mutations in the cii gene, one of a few sites in lambda at which such mutations arise, were located genetically and physically with overlapping deletions. dna sequence analysis of the deletion segment containing the x+ c mutations showed that two independent x+ c mutations arose by the same a-t to t-a transversion. presumably ...1978282634
genetic inversion in the formation of an hfr strain from a temperature-sensitive f' gal hfr strain (pb15) that carries a duplicated copy of the galactose operon genes flanking the integrated sex factor is unusually stable since it does not show excision of the repeated deoxyribonucleic acid segment. the right-hand galactose operon is in the normal orientation. deletion mutations that eliminate the right-hand galactose genes, the sex factor, and some of the left-hand operon have been isolated. mutants believed to have their left-hand galactose operon inverted were able to be indu ...1977318642
isolation and characterization of lambda pleu the escherichia coli lysogen hfrh73 described by shimada et al. (1973), none of the enzymes coded for by the leucine operon is synthesized due to an insertion of phage lambda into cistron leua. the orientation of lambda in the chromosome is ara leudcb lambda jan leua. after heat induction of the lysogen, plaque-forming transducing phages of two types are formed at low frequency. one type (e.g., lambda pleu9) transduces leud, leuc, and leub strains to prototrophy. the other type (e.g., lambda ...1977320178
isolation and characterization of conditional-lethal rho mutants of escherichia coli.temperature-sensitive nita (rho) mutants of e. coli were isolated; one of them was characterized as an amber mutant. these strains show the nit phenotype (transcription of phage lambda dna independent of the n gene) at low temperatures and are inviable at high temperatures. the mutated sites appear to be between cya and mete on the chromosome. temperature-sensitive nita bacteria not only permit leftward transcription of the lambda genome at a high rate in the absence of the lambda n protein, but ...1977322147
isolation and characterization of plaque-forming lambdadnaz+ transducing bacteriophages.the escherichia coli dnaz gene, a deoxyribonucleic acid (dna) polymerization gene, is located 1.2 min counterclockwise from pure, at approximately min 10.5 on the e. coli map. from a lysogen with lamdaci857 integrated at a secondary attachment site near pure, transducing phages (lambdadnas+) that transduced a dnazts (lambda+) recipient to temperature insensitivity (ts+) were discovered. three different plaque-forming transducing phages were isolated from seven primary heterogenotes. genetic test ...1977323235
transcription in vitro of bacteriophage lambda 4s rna: studies on termination and rho protein.when bacteriophage lambdapga18 dna is transcribed in a purified in vitro system by e. coli rna polymerase (nucleoside triphosphate: rna nucleotidyl-transferase, ec, several major transcripts are synthesized. we have investigated transcriptional termination of one of these transcripts, the 4s, or "oop" rna. analysis by two-dimensional "fingerprinting" of t1 oligonucleotides reveals that transcription of the 4s rna terminates at a specific site on the lambdapga18 dna template, t-l with an ...1977325526
genetic characterization of plasmid formation by n- mutants of bacteriophage lambda. 1977325881
analysis of the phase variation in lambda reduced immunity lysogens.two distinct phases characterized by different levels of immunity that appear in some e. coli strains lysogenic for reduced immunity mutants of bacteriophage lambda are identified as single and tandem double lysogens respectively on the basis of dna-dna hybridization experiments and the requirement of the phage xis function for the transition from a single to a double, and of the host reca function for the transition from a double to a single lysogen (in a xis- condition). rim lysogens with a fu ...1977327265
a specialized transducing lambda phage carrying the escherichia coli genes for phenylalanyl-trna synthetase.a lambda phage has been isolated which specifically transduces the escherichia coli phes and phet genes coding for the alpha and beta subunits of the phenylalanyl-trna synthetase (prs). this phage transduces with high frequency (i) several temperature-sensitive prs mutants to thermoresistance and (ii) a p-fluorophenylalanine resistant prs mutant to sensitivity against this amino-acid analog. the in vitro prs activities of such lysogens suggest that the alpha and beta subunits coded by the transd ...1977327276
[the action of selected herbicides on bacteriophages and escherichia coli (author's transl)].the effect of eight herbicides on the multiplication of bacteria and bacteriophages was tested with escherichia coli, strains w1665f+ and c600, and with the rna-phage m12 and the dna-phage lambda in turbidimetric investigations and one-step growth experiments. e. coli is inhibited by seven of the herbicides investigated in concentrations of 10(-3)m, partly of 10(-4)m, too, and is promoted by some compounds in weaker concentrations. naphthylacetic acid, (nes) largely independent of its concentrat ...1977327728
the mechanism of action of nitro-heterocyclic antimicrobial drugs. primary target of 1-methyl-2-nitro-5-vinylimidazole is dna.the antimicrobial drug 1-methyl-2-nitro-5-vinylimidazole (mev) preferentially blocked dna synthesis, was mutagenic and induced coliphage lambda in escherichia coli. the antibacterial effects of mev are the consequences of repairable damage to dna, as shown by hypersensitivity of reca and uvr strains to mev and related drugs, stimulation by mev of dna turnover which was dependent on the product of the uvra gene, and the presence of cross-links in dna from mev-treated bacteria.1977330809
kinetics of induction of error-prone repair of bacteriophage lambda by temperature shift in an escherichia coli dnab mutant.preincubation at 42 degrees, before infection at permissive temperature by phage lambda, of an escherichia coli dnab mutant, provokes a significant increase in survival and mutagenesis of ultraviolet irradiated phage as well as mutagenesis of untreated phage. similarly to uv irradiation and many chemical mutagens, the inhibition of dna synthesis by temperature shift of this dnab mutant induces sos repair. this work shows that replication blockage in bacterial dna is not only mutagenic for bacter ...1977337133
in vitro packaging of a lambda dam vector containing ecori dna fragments of escherichia coli and phage this report we describe a coliphage lambda vector system for cloning endo r. ecori dna fragments. this system differs significantly from those previously described in two ways. first, restricted and ligated dna is encapsidated in vitro. second, with increasing lambda dna size in the range 78 to 100% that of wild-type, the efficiency of dna encapsidation into infectious phage particles markedly increases. for lambda wild-type dna the efficiency of in vitro packaging (10(6) to 10(7) plaques pro ...1977338419
maximizing gene expression on a plasmid using recombination in vitro.recombination in vitro has been used to place one or more copies of a strong promoter, the lac promoter, at varying distances from the cl (repressor) gene of bacteriophage lambda on the e. coli plasmid pmb9. in all constructions, lambda repressor synthesis is driven wholly or predominantly by the inserted lac promoter. one of our fusions directs the synthesis of very high levels of lambda repressor. in this case, the fused dna encodes a ribosome binding site which is a "hybrid" of lambda and lac ...1978340049
the thirteenth colworth medal lecture: the construction in vitro and exploitation of transducing derivatives of bacteriophage lambda. 1977340298
initiation of lambda dna replication in vitro promoted by isolated p-gene product.the product of the p gene of bacteriophage lambda was isolated from heat-induced lambda-lysogenic escherichia coli cells. it was found to bind to dna, to be devoid of nuclease activity acting on double-stranded lambda dna and of nicking/closing activity. initiation of lambda dna replication promoted by the p-gene product in a complementation assay in vitro was sensitive to rifampicin. sedimentation analysis of the products and their hybridization to separated lambda dna strands indicate that lam ...1978342246
genes for the hook-basal body proteins of the flagellar apparatus in escherichia coli.of the more than 30 genes required for flagellar function, 6 are located between pyrc and ptsg on the escherichia coli genetic man. this cluster of genes is called flagellar region i. four-point transductional crosses were used to establish the position and order of the region i flagellar genes with respect to the outside markers ptsg and pyrc. bacteriophage lambda-e. coli hybrids that contained most of the genes necessary for flagellar formation were constructed. the properties of specific hybr ...1978350831
transfection of escherichia coli spheroplasts: infectious lambda prophage dna.high mol. wt. dna was extracted from escherichia coli lambda lysogens and was shown to be infectious. its infectivity was due to prophage dna integrated into the host chromosome rather than to dna released from mature phage particles, as established by the following criteria: the titre of infectious dna exceeded by 100-fold the titre of infectious units present before dna extraction; mild shear selectively reduced prophage dna infectivity to 2% of the unsheared dna while lambda phage dna infecti ...1978351139
correlation between uv dose requirement for lambda bacteriophage induction and lambda repressor concentration.escherichia coli k-12 wild type and a uvra mutant derivative were used to construct isogenic strains bearing one, two, three, or more phage lambda ci genomes and containing increasing concentration of lambda repressor as measured by in vitro operator dna-binding assays. the survival and phage induction in response to uv irradiation were determined. in both strains, dose-response relationships were obtained as a function of the cellular repressor concentration. the uvra lysogens required one-tent ...1978353300
bacteriophage lambda carrying the escherichia coli chromosomal region of the replication origin.a transducing phage lambdaasn was isolated. the late gene region of its genome was found to have been substituted by an escherichia coli chromosomal segment containing the genes bgir, bgic, glms, unca, and asn. restriction endonuclease cleavage mapping and electron microscopic analysis of the lambdaasn dna revealed that the size of the bacterial segment is approximately 1.75 x 10(7) daltons, corresponding to about 26.4 kilobases. the circular dna of lambdaasn was digested with restriction endonu ...1978368808
inactivation of bacteriophage lambda by combined x-ray and u.v.-light exposure.extracellular phage lambda has been successively exposed to x-rays and u.v. light. the plaque-forming ability of the irradiated phages was determined on host cells with different repair capacities. no change in sensitivity was found with a pre-treatment of one type of radiation to lethal damage inflicted by the other. this indicates that a prerequisite for an interaction of different types of radiation is either an active metabolism or repair process occurring during the two radiation exposures.1979381227
transduction of bacteriophage lambda by bacteriophage t1.when bacteriophage t1 was grown on bacteriophage lambda-lysogenic cells, phenotypically mixed particles were formed which had the serum sensitivity, host range, and density of t1 but which gave rise to lambda phage. t1 packaged lambda genomes more efficiently both when the length of the prophage was less than that of wild-type lambda and when the host cell was polylysogenic. expression of the red genes of lambda or the rece system of escherichia coli during t1 growth enhanced pickup of lambda by ...1979381686
in vitro transcription by wheat germ ribonucleic acid polymerase ii: effects of heparin and role of template integrity.double-stranded deoxyribonucleic acid (dna) from bacteriophage lambda is a good template for wheat germ dna-dependent ribonucleic acid (rna) polymerase ii. we delineated conditions for obtaining maximum polymerase activity using as template both the relatively intact dna extracted from the the lambda phage and dna into which single-strand nicks have been introduced by deoxyribonuclease (dnase) i digestion. the deliberate introduction of nicks produces a modest increase in transcription. the nacl ...1979387073
recombination of bacteriophage phi x174 by the red function of bacteriophage lambda.recombination of bacteriophage phi x174 was effectively promoted when the red function of lambda was supplied by either co-infection with lambda or induction of lambda lysogens. mutations in red alpha and red beta genes of lambda abolished recombination nearly completely, whereas a mutation in gam gene reduced it only slightly. the red-promoted recombination of phi x174 occurred in reca, recb, and pola mutants as well as in wild-type strains of escherichia coli. it was further stimulated when ph ...1979430603
isolation and properties of escherichia coli mutants which are nonpermissive for the growth of phage selecting survivors of lambda phage infection, mutants of escherichia coli k12 that block reproduction cycle of the phage have been isolated. fourteen of these phage-tolerant mutants (lam mutants) were chosen and characterized biochemically and genetically. it was shown that these mutants were tolerant to infection by all the lambdoid phages, except for few cases, but they were susceptible to infection by a non-lambdoid temperate phage (phi299), p1 or t phages. the mutants can be divided into ...1975772257
morphogenesis of bacteriophage lambda: electron microscopy of thin sections. 1976781268
transposition and fusion of the lac genes to selected promoters in escherichia coli using bacteriophage lambda and mu. 1976781293
lethal lysogenization by coliphage lambda. 1976782020
specialized transduction of colicin e1 dna in escherichia coli k-12.genetic studies were made on e. coli k-12 tm96, which carries recombinant molecules constructed by in vitro combination of colicin e1 dna and a dna fragment of e. coli for guanine synthesis derived from transducing phage. the recombinant molecules existed as stable plasmids within the cell and contained genes for colicin e1 immunity and the guaa enzyme (xanthosine 5'-monophosphate aminase) together with a part of the lambda genome, r through j: (r-a-f-j)+. a block of the lambda genome, int throu ...1976787989
maturation of a single lambda phage particle from a dimeric circular lambda dna. 1976790755
a transducing bacteriophage lambda carrying the structural gene for elongation factor ts.a specialized transducing bacteriophage lambdadpolcdap d-9 has been isolated that carries the structural gene for ef-ts1 (tsf). the presence of ef-ts among the proteins synthesized under the direction of this phage in uvl-inactivated cells has been detected by two-dimensional gel electrophoresis and has been verified by antibody precipitation. in an induced lysogen of this phage the relative rate of synthesis of ef-ts is increased 4-fold. evidence is presented which suggest that the structural g ...1976792684
suppressible mutations in the cro gene of bacteriophage lambda. 1976795138
physicochomecial studies on interactions between dna and rna polymerase. isolation and mapping of a t7 dna fragment containing the early promoters for escherichia coli rna polymerase.the cleavage sites in the early promoter region of coliphage t7 have been mapped for four restriction enzymes. they are, from the left end in base pairs, 1100 and 740 for hinf; 680, 320, 530, 240, 77, and 67 for hind ii; 620 and 530 for hpa ii; 790 for alu i. the nucleotide sequence between the hind ii site at 680 base pairs from the left end and the hinf site at 740 base pairs from the left end has been determined, from which the start point of the promoter a3 is located at 720 base pairs from ...1976795461
[new mutations in the early stage of phage lambda]. 1976798250
[substrate of a uv-induced repair system providing for w-reactivation of lambda phage].escherichia coli uvra, pola and uvrd cells carrying non-uv-inducible prophage lambdac1857ind- were infected with 3h-thymidine labelled homoimmune phage lambdac1857, and the effect of uv-irradiation of super-infecting phage and lysogenic bacterial cells on the content of intracellular covalently-closed lambda dna circles (cccdna) and pyrimidine dimer content in lambda dna are studied. uv-irradiation of host cells results in two-fold increase of relative content of cccdna of uv-irradiated phage la ...19761001892
location in bacteriophage lamdba dna of cleavage sites of the site-specific endonuclease from bacillus amyloliquefaciens h.the sites in escherichia coli bacteriophage lambda dna cleaved by the site-specific endonuclease isolated from bacillus amyloliquefaciens h (bami) are found to be at 0.114, 0.466, 0.580, 0.713, and 0.861 lambda units. the sites were located by analysis of the products of digestion of lambda dna and lambda-ara transducing phage dna, and verified by double digestion with bami and ecori.19761083914
morphogenesis of the tail of bacteriophage lambda. ii. in vitro formation and properties of phage particles with extra long tails. 19751089336
proteolytic cleavage of bacteriophage lambda repressor in induction.the bacteriophage lambda repressor, a protein that maintains the lysogenic state of a bacterium containing a lambda prophage, is cleaved when the lysogen is induced by mitomycin c or ultraviolet light. this cleavage does not occur when induction is prevented by mutational alteration either of the phage repressor or of the host reca gene product. proteolytic cleavage may be the primary mechanism of repressor inactivation in this induction pathway, or it may follow a different event which causes t ...19751090931
mutations simultaneously affecting endonuclease ii and exonuclease iii in escherichia coli.we studied mutants of e. coli originally identified as being deficient in either endonuclease ii (deoxyribonucleate oligonucleotidohydrolase, ec or exonuclease iii [deoxyribonucleate (double-stranded) 5'-nucleotidohydrolase, ec] activity. twelve independently derived mutants were tested, including three new endonuclease ii mutants. deficiency of one enzyme was always accompanied by deficiency of the other. furthermore, temperature-sensitivity of one activity was always accompa ...19751091930
mismatch repair in heteroduplex dna.dna with base pair mismatches was prepared by annealing mixtures of genetically marked dna from bacteriophage lambda. this heteroduplex dna was used to transfect bacteria under conditions minimizing recombination. genetic analysis of the progeny phages indicates that: (i) mismatch repair occurs, usually giving rise to a dna molecule with one chain with the genotype arising from repair and one parental chain. (ii) the frequency of repair of a given mismatch to wild type depends on the marker, ran ...19751094458
constitutive integrative recombination by bacteriophage lambda. 19751094679
processing of bacteriophage lambda dna during its assembly into heads. 19751102699
studies on the cleavage of bacteriophage lambda dna with ecori restriction endonuclease. 19751102702
bacteriophage lambda induction causes increased production of e. coli lysine transfer rna. 19751103738
phage lambda receptor chromosomes for dna fragments made with restriction endonuclease iii of haemophilus influenzae and restriction endonuclease i of escherichia coli. 19751104875
reversible interaction between coliphage lambda and its receptor protein. 19751107562
the yield of radiation-induced dna single-strand breaks in escherichia coli and superinfecting phage lambda at different dose rates. repair of strand breaks in different buffers. 1976794917
induction of sigma factor synthesis in escherichia coli by the n gene product of bacteriophage lambda.thermoinduction of cells of e. coli carrying prophage lambdaci857 within the bfe gene brings about not only "escape synthesis" of core subunits of the dna-dependent rna polymerase (rna nucleotidyltransferase, nucleosidetriphosphate:rna nucleotidyltransferase, ec 2-7-7-6), but also a striking stimulation of sigma factor synthesis. the latter phenomenon, termed sigma induction, is generally observed after lambda phage infection or prophage induction. a series of experiments with various bacterial ...1976794877
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 ...19921310522
[repair of recombinant plasmids].comparative analysis of uv-sensitivity was carried out on plasmids of various molecular weight. recombinant plasmids containing fragments of prokaryotic dna (e. coli, phage lambda) are repaired in e. coli cells more effectively than those containing eukaryotic dna fragments. it was also shown that uv-sensitivity of recombinant plasmids is independent of their molecular weight provided that the active repair process in fact occurs. uv-sensitivity was strongly proporational to dna size only when e ...1979398000
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.19921327960
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 ...19921327972
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 ...19921336178
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 ...19901368559
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 ...19921373849
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 ...19921374802
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 ...19921385814
construction of coliphage lambda charon vectors with bamh1 cloning sites. 1980. 19921422018
physical characterisation of the "rac prophage" in e. coli k12.we confirm the hypothesis of low (1973) that many e. coli k12 strains contain a prophage (the rac prophage) located a few minutes clockwise of the trp operon on the genetic map. we have used restriction endonucleases and 32p-labelled probes to construct a physical map of this prophage. some e. coli k12 strains, including ab1157, have lost the entire prophage, apparently by a specific deletion. this is consistent with prophage excision by site-specific recombination. lambda reverse (lambda rev) p ...1979390313
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 ...19921435879
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 ...19921439823
interaction of bacteriophage k10 with its receptor, the lamb protein of escherichia coli.the lamb protein of escherichia coli was initially recognized as the receptor for bacteriophage lambda. it is now shown also to constitute the receptor for phage k10. the lamb protein interacts with phage k10 in vitro, but this interaction does not lead to phage inactivation. most lambda-resistant labb mutants are also resistant to k10, and vice versa. however, a significant proportion of the mutants resistant to one of the phages is sensitive to the other. nineteen k10-resistant lambda-sensitiv ...1979387746
[molecular cloning of the structural genes of the escherichia coli deo-operon in plasmid rsf2124].a specialized phage lambda ddeo carrying the deo operon of escherichia coli is analyzed by exposing the dna to the specific restriction endonucleases ecori and bamhi. using the lambda ddeo dna fragment, obtained by digestion with bamhi and plasmid rsf2124 as a vehicle, the hybrid plasmid pam1 carrying all the genes of the deo operon is constructed and cloned in e. coli cells. it is shown that the activity of thymidine phosphorylase in the strain am061, which contains hybrid plasmid pam1 is 30-fo ...1978363516
restoration of polarity by n-deficiency in lambda phage containing a translocated trp operon segment.when translation of trp mrna is terminated by a nonsense codon or by antibiotics like chloramphenicol, the amount of the mrna distal to the blocked ribosomes is found at much lower levels ("polarity"). polarity is alleviated when the trp mrna is formed as part of a long transcript from the phage lambda promoter pl (segawa and imamoto, 1974; franklin, 1974); but the relief of polarity is itself largely dependent on the lambda protein n. in a phage that joins the trp operon segment (trpd, c, b a) ...1978345080
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 ...19921533056
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 ...19921533631
insertion sequence is2 associated with int-constitutive mutants of bacteriophage lambda.we have examined mutations in bacteriophage lambda called int-c, which confer elevated constitutive expression on the int gene for prophage integration. one class of mutations, which map between the b538 and bio386 endpoints, does not appear to be associated with any major chromosomal modification, whereas the second class has the is2 insertion sequence in orientation ii within the region between gene int and the b538 endpoint, all int-c mutations are within gene xis, with the possible exception ...1977344135
induction of error-prone repair as a consequence of dna ligase deficiency in escherichia coli.dna ligase deficiency is shown to induce generalized mutator activity in e. coli. this mutator activity is unaffected by 3 mug/ml of chloramphenicol but is abolished both in lig-reca double mutants and by incubation with 20 mug/ml of chloramphenicol. dna ligase deficiency is also shown to reactivate ultraviolet light-irradiated phage lambda and t7 and to increase both spontaneous and ultraviolet light-induced mutagenesis in phage lambda, all of which are abolished in lig-reca strains. interactio ...19751105587
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 ...19921610363
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 ...19911646785
the biological activity of bacteriophage dna, prepared by the cationic detergent dilution technique.the preparation of phage lambda dna infecting e. coli k 12 with cationic detergent is described. this dna infects e. coli spheroblasts with the same efficiency as dna prepared by phenol methods.19751093138
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 ...19921316868
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 ...19911657185
coupling of lac mrna transcription to translation in escherichia coli cell an extract containing all the components for lac gene expression except washed ribosomes, lac mrna formation was increased 4- to 6-fold by the addition of washed ribosomes. the formation of beta-galactosidase mrna and enzyme showed very different dependency on added ribosomes. enzyme was formed in proportion to the number of ribosomes added, whereas 10% of the standard level of ribosomes promoted full levels of transcription. consistent with their action in vivo, chloramphenicol and erythromy ...1978415305
temperature-inducible gene expression in bacillus subtilis mediated by the ci857-encoded repressor of bacteriophage 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 ...19901699846
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 ...19921730025
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 ...19921730643
hu and integration host factor function as auxiliary proteins in cleavage of phage lambda cohesive ends by 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 ...19911825651
in vitro synthesis of beta-galactosidase with ilv-lac fusion deoxyribonucleic acid as in vitro protein-synthesizing system has been developed to study the mechanism of induction of ilvc gene in escherichia coli strain k-12. deoxyribonucleic acid (dna) from a lambda phage carrying an ilvc-lac fusion was employed as a template for the in vitro synthesis of beta-galactosidase under the control of the ilvc promoter. the use of this template allowed an investigation of the components required for induction of the ilvc gene and the kinetics of the induction. the in vitro synthesis o ...1977411784
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 ...19911827523
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 ...19921452017
cole1 plasmid incompatibility: localization and analysis of mutations affecting incompatibility.deletion mutants of plasmid cole1 that involve the replication origin and adjacent regions of the plasmid have been studied to determine the mechanism by which those mutations affect the expression of plasmid incompatibility. it was observed that (i) a region of cole1 that is involved in the expression of plasmid incompatibility lies between base pairs -185 and -684; (ii) the integrity of at least part of the region of cole1 dna between base pairs -185 and -572 is essential for the expression of ...1979378980
repressor and int synthesis of bacteriophage lambda in the e. coli host mutant er437.analysis of lambda phage infection of the host mutant er437 by sds polyacrylamide gel electrophoresis and autoradiography has revealed altered expression of repressor and integration function (int). we show that in this host int as well as repressor synthesis is not dependent upon the lambdaciii gene product in the usual manner, nor is their synthesis turned off in the normal way.1977340886
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 ...19911829453
dna replication--bacteriophage lambda. 1977340149
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 ...19911830578
specific hydrolysis of the cohesive ends of bacteriophage lambda dna by three single strand-specific nucleases.procedures have been worked out for aspergillus nuclease s1 and mung been nuclease to quantitatively cleave off both of the 12-nucleotide long, single-stranded cohesive ends of lambdadna. this cleavage is indicated by the almost complete elimination of the repair incorporation of radioactive nucleotides by dna polymerase into the digested dna. with s1 nuclease, cleavage was complete at 10 degrees as well as at 30 degrees. under the conditions for quantitative cleavage of the single-stranded regi ...19751141222
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