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replication region fragments cloned from flac+ are identical to ecori fragment f5 of f.the replication region fragments from flac(+) cloned in plasmids psc138 and pml31 are identical with each other and with ecori fragment 5 of plasmid f.1976956131
host-parasite interactions: recent developments in the genetics of abortive phage infections.recent progress towards the elucidation of the mechanisms of phage exclusion systems that lead to abortive infections is described. abortive infections constitute model systems for the study of host-parasite interactions, but until the 1980s such studies were confined largely to descriptions of a plethora of physiological dysfunctions. the use of cloning vectors containing the genes involved in the exclusion phenomena have helped to reduce the complexity of the abortive infection while maintaini ...19911831658
the hok killer gene family in gram-negative bacteria.the seven members of the hok killer gene family in gram-negative bacteria are described here. the members of this gene family have been sequenced and include hok/sok from plasmid r1, flm and srnb from plasmid f, pnd from plasmids r483 and r16, and gef and relf, which are located on the escherichia coli chromosome. the killer proteins encoded by these loci are highly toxic polypeptides of 50 to 52 amino acids. the proteins kill the cells from the inside by interfering with a vital function in the ...19902101633
the ssb gene of plasmid colib-p9.the inci1 plasmid colib-p9 was found to carry a single-stranded dna-binding (ssb) protein gene (ssb) that maps about 11 kilobase pairs from the origin of transfer in the region transferred early during bacterial conjugation. the cloned gene was able to suppress the uv and temperature sensitivity of an ssb-1 strain of escherichia coli k-12. the nucleotide sequence of the colib ssb gene was determined, giving a predicted molecular weight of 19,110 for the ssb protein. sequence data show that colib ...19892651402
control of the ccd operon in plasmid f.the f sex factor plasmid of escherichia coli contains a pair of genes, ccda and ccdb, whose protein gene products are involved in an unusual feature of plasmid maintenance. the ccdb protein is a cytotoxin that becomes activated when the f plasmid is lost, thereby killing the f- segregant cells. in f+ cells, the ccda protein protects against the lethal effects of ccdb. in the present study we show that ccda and ccdb expressions are negatively autoregulated at the level of transcription. genetic s ...19892651399
analysis of escherichia coli k12 f factor transfer genes: traq, trba, and trbb.the genes that encode the transfer properties of plasmid f, the fertility factor of escherichia coli k12, are known to be clustered over a large, 33.3-kb segment of f dna. as the central segment of the transfer region has not previously been well characterized, we constructed a detailed restriction map of the large f ecori dna fragment, fl, and isolated a series of plasmid derivatives that carry various overlapping segments of this f tra operon dna. we also analyzed the protein products of those ...19872827204
expression of virulence and antibiotic resistance in an escherichia coli transconjugant carrying a large plasmid pcat120 of shigella dysenteriae type i and its spontaneous fragmentations.the role of a 120-kb plasmid in relation to virulence and drug resistance factor in shigella dysenteriae was studied. for characterization of plasmids, the mating system is a useful and efficient means of transferring both large and small plasmids to a new host. the conjugative transfer of a 120-kb (pcat120) ampicillin-resistant plasmid of s. dysenteriae to e. coli k-12 was not successful. introduction of an e. coli fertility factor plasmid f, did not help to mobilize the plasmid. low transfer f ...19911823646
escherichia coli minichromosomes: random segregation and absence of copy number control.minichromosomes, i.e. plasmids that can replicate from an integrated oric, have been puzzling because of their high copy numbers compared to that of the chromosomal oric, their lack of incompatibility with the chromosome and their high loss frequencies. using single cell resistance to tetracycline or ampicillin as an indicator of copy number we followed the development of minichromosome distributions in escherichia coli cells transformed with minichromosomes and then allowed to grow towards the ...19902213882
the plasmid-maintenance functions of the p7 prophage.the region responsible for the maintenance of the prophage of bacteriophage p7 as a stable, unit-copy plasmid was isolated in a lambda att vector which lysogenizes escherichia coli as a stable unit-copy plasmid under the control of the p7 replication origin. the p7 plasmid-maintenance region was shown to consist of adjacent replication and partition regions capable of functioning independently. the isolated replication region could support plasmid maintenance but the resulting plasmids were high ...19872827210
the tram protein of the conjugative plasmid f binds to the origin of transfer of the f and cole1 plasmids.the gene encoding the tram protein of the conjugative plasmid f was cloned, overexpressed and the gene product was purified. the tram protein was found in the cytoplasm of cells carrying the f plasmid with a smaller amount in the inner membrane. dnase i footprinting experiments showed that the purified protein protects three regions in the f orit locus with different affinity for the upper and lower strands of dna. a 15-nucleotide motif was identified within the protected regions that represente ...19921479887
integrative suppression of dnaa(ts) mutations mediated by plasmid f in escherichia coli is a dnaa-dependent process.the thermosensitivity of dnaa(ts) mutations can be suppressed by integration of plasmid f (integrative suppression). in the light of the recent finding that f requires dnaa protein for both establishment and maintenance, integrative suppression of 11 dnaa(ts) mutations by a mini-f, pml31, integrated near oric was examined. the plating efficiency of integratively suppressed strains was dnaa(ts) allele-dependent and medium-dependent. the initiation capability of suppressed dnaa(ts) strains lacking ...19872830456
nucleotide sequence of the promoter-distal region of the tra operon of plasmid r100, including trai (dna helicase i) and trad genes.the nucleotide sequence of the promoter-distal region of the tra operon of r100 was determined. there are five open reading frames in the region between trat and fino, and their protein products were identified. nucleotide sequences of plasmid f corresponding to the junction regions among the open reading frames seen in r100 were also determined. comparison of these nucleotide sequences revealed strong homology in the regions containing trad, trai and an open reading frame (named orfd). the trad ...19902164585
nucleotide sequence of the leading region adjacent to the origin of transfer on plasmid f and its conservation among conjugative plasmids.the leading region of the escherichia coli k12 f plasmid is the first segment of dna to be transferred into the recipient cell during conjugal transfer. we report the nucleotide sequence of the 64.20-66.77f portion of the leading region immediately adjacent to the origin of transfer, orit. the 2582 bp region encodes three open reading frames, orf95, orf169 and orf273; the product of orf273, is equivalent in size and map location to the 35 kda protein, 6d, previously described (cram et al. 1984). ...19892693941
the 41 carboxy-terminal residues of the minif plasmid ccda protein are sufficient to antagonize the killer activity of the ccdb protein.the ccd operon of plasmid f encodes two genes, ccda and ccdb, which contribute to the high stability of the plasmid by post-segregational killing of plasmid-free bacteria. the ccdb protein is lethal to bacteria and the ccda protein is an antagonist of this lethal action. a 520 bp fragment containing the terminal part of the ccda gene and the entire ccdb gene of plasmid f was cloned downstream of the tac promoter. although the ccdb protein was expressed from this fragment, no killing of host bact ...19912034222
site-specific proteolysis of mini-f plasmid replication protein repe destroys initiator function and generates an incompatibility substance.plasmid f replication is controlled by a plasmid-specified rep protein with both autorepressor and initiator functions. the mechanism by which these two functions of a rep protein are balanced to achieve stable replication is unknown; however, we speculated in prior work that rep protein modification could be involved. we report here that naturally proteolyzed f repe protein has been detected and characterized. the processed molecule lost the first 17 n-terminal aminoacyl residues and initiator ...19921569028
cloning and stable maintenance of 300-kilobase-pair fragments of human dna in escherichia coli using an f-factor-based vector.a bacterial cloning system for mapping and analysis of complex genomes has been developed. the bac system (for bacterial artificial chromosome) is based on escherichia coli and its single-copy plasmid f factor. it is capable of maintaining human genomic dna fragments of greater than 300 kilobase pairs. individual clones of human dna appear to be maintained with a high degree of structural stability in the host, even after 100 generations of serial growth. because of high cloning efficiency, easy ...19921528894
location of enzymatic and dna-binding domains on e. coli protease la.escherichia coli protease la is an atp-dependent enzyme that has a dna-binding site. the locations of the enzymatic and dna-binding sites are not known. we report that a 75-residue segment at the carboxy-terminus of the protease la is similar to part of bacillus licheniformis beta-lactamase, a serine enzyme. the comparison score is 8.2 standard deviations higher than that obtained with 10,000 comparisons of randomized sequences of these segments. the probability of obtaining such a score by chan ...19892647519
the f plasmid ccd autorepressor is a complex of ccda and ccdb proteins.the ccd operon of plasmid f produces three proteins, ccda, ccdb, and repd. prior research has established that the operon is autorepressed and that at least ccdb, but not repd, is required for autorepression. a role for ccda in autorepression was suggested but not clearly shown. we now present a series of biochemical experiments which show that both ccda and ccdb proteins are required for maximal formation of protein-ccd operator complexes. we also show that ccda and ccdb are present in a comple ...19892615761
sequence alterations affecting f plasmid transfer gene expression: a conjugation system dependent on transcription by the rna polymerase of phage t7.we constructed derivatives of the escherichia coli conjugative plasmid f that carry altered sequences in place of the major transfer operon promoter, py. replacement of py with a promoter-deficient sequence resulted in a transfer-deficient, f-pilus-specific phage-resistant plasmid (pox38-tra701) that could still express traj and trat; tray, f-pilin, trad, and trai were not detectable on western blots. on a second plasmid (pox38-tra715) we replaced py with a phage t7 late promoter sequence. in ho ...19921479888
conjugal transfer of cloning vectors derived from cole1.the transfer properties of five cloning vectors derived from cole1 were studied. two of the vectors (psf2124 and pgm706) behaved like wild type cole1 in that they could be transferred efficiently in the presence of the conjugative plasmid f. the mobilization of the remaining three vectors (pmb9, pbr313 and pbr322) by f was barely detectable. the transfer defect in pbr313 and pbr322 could be complemented by colk when r64drd11, but not f, was used as the conjugative plasmid. the transferred plasmi ...1978363522
restriction endonuclease mapping and mutagenesis of the f sex factor replication region.the plasmids psc138 and pml31 each contain the ecori-generated f5 replicator fragment of the conjugative plasmid f in addition to an ecori fragment encoding antibiotic resistance: ampicillin resistance derived from staphylococcus aureus in psc138 and kanamycin resistance from escherichia coli in pml31. we have mapped one hindiii and two bamhi restriction sites in the f5 region of these plasmids and one hindiii site in the antibiotic resistance region of each plasmid. the hindiii site in the km r ...1977327274
interaction of the expression of two membrane genes, acra and plsa, in escherichia coli k-12.the mutation acra1, leading to acriflavine sensitivity through disorganization of the plasma membrane, is located between proc and pure on the escherichia coli k-12 chromosome. gene plsa has been reported to determine biosynthesis of membrane phospholipid and to be located very near acra (1). genes acra and plsa fall into different cistrons and are arranged in the order proc-acra-plasa-pure. the genes were shown to interact with each other. introduction of acra mutation into a plsa temperature-s ...19751097404
nucleotide sequences of the r1-19 plasmid transfer genes tram, finp, traj, and tray and the trayz promoter.the complete nucleotide sequences of the r1 drd-19 (r1-19) plasmid transfer genes tram, finp, traj, and tray and the region encoding the trayz promoter were determined. the tram protein from r1-19 was similar to the 127-amino-acid tram product from the conjugative plasmid f; only 28 residues were not identical. finp, a negative regulatory element of the traj gene, contained a 12-base-pair inverted repeat identical to that found in the f plasmid, but differed in the 7 base pairs found between the ...19863009392
requirement of the escherichia coli dnaa gene product for plasmid f maintenance.there are dnaa protein-binding sites in at least one f origin of replication, and only potentially leaky dnaa(ts) mutations had ever been used in previous studies indicating that f replication was independent of the dnaa gene product. here we show that an escherichia coli dnaa::tn10 host which does not make a dnaa gene product cannot sustain autonomous or integrated f plasmid maintenance.19863020005
repressor gene fino in plasmids r100 and f: constitutive transfer of plasmid f is caused by insertion of is3 into f fino.fertility factor f confers bacterial conjugation, a process which involves at least 20 tra genes. resistance plasmids such as r100, r6-5, and r1 have homology with f in the tra region. conjugal transfer of these plasmids is, however, repressed, while transfer of f is constitutive. repression of r transfer is due to the existence of the two genes, called fino and finp; constitutive transfer of f is believed to be due to a lack of fino in f. in this paper, we report the identification and dna sequ ...19873027040
structural and functional comparison between the stability systems pard of plasmid r1 and ccd of plasmid f.the stability determined by the systems pard of plasmid r1 and ccd of plasmid f is due to the concerted action of two proteins, a cytotoxin and an antagonist of this function. in this paper we report that ccda and kis proteins, the antagonists of the ccd and pard systems respectively, share significant sequence homologies at both ends. in kis, these regions seem to correspond to two different domains. despite the structural similarities, kis and ccda are not interchangeable. in addition we have ...19912017133
genetic analysis of an invasion region by use of a tn3-lac transposon and identification of a second positive regulator gene, inve, for cell invasion of shigella sonnei: significant homology of inve with parb of plasmid p1.we have previously cloned two distinct regions of the shigella sonnei form i plasmid pss120, a 37-kilobase-pair dna region and a virf region, which were found to be essential for cell invasion in escherichia coli k-12 (j. kato, k. ito, a. nakamura, and h. watanabe, infect. immun. 57:1391-1398, 1989). the 37-kilobase-pair dna region was randomly inserted by use of transposon tn3-lac. at least eight genes were found to be located within the region, as determined by analysis of tn3-lac-generated la ...19901688841
fertility repression of f-like conjugative plasmids: physical mapping of the r6--5 fino and finp cistrons and identification of the fino protein.the locations of the fertility inhibition genes fino and finp of the f-like conjugative multiple antibiotic-resistance plasmid r6-5 have been determined. as found previously for that of the fertility plasmid f, the finp gene of r6-5 is located close to the origin of dna transfer, orit, and to the promoter-proximal segment of the tra operon. thus, finp is close to the site of action of the finop fertility inhibition system. in contrast, the fino gene is located on the other side of the tra operon ...1978366604
stability and replication control of escherichia coli minichromosomes.a stabilized minichromosome--a plasmid replicating from the chromosomal origin oric--was constructed by cloning the sopa,b,c, genes from plasmid f. this minichromosome had a loss frequency of less than 10(-3), while that of the nonstabilized parental plasmid was 2 x 10(-2) to 4 x 10(-2). both minichromosomes had the same average copy number per chromosomal origin, and the copy numbers were constant over an eightfold range of growth rates. different mutations in the mioc gene and promoter, from w ...19873294807
the rifampicin-inducible genes srnb from f and pnd from r483 are regulated by antisense rnas and mediate plasmid maintenance by killing of plasmid-free segregants.the gene systems srnb of plasmid f and pnd of plasmid r483 were discovered because of their induction by rifampicin. induction caused membrane damage, rnase i influx, degradation of stable rna and, consequently, cell killing. we show here that the srnb and pnd systems mediate efficient stabilization of a mini-r1 test-plasmid. we also show that the killer genes srnb' and pnda are regulated by antisense rnas, and that the srnc- and pndb-encoded antisense rnas, denoted srnc- and pndb-rnas, are unst ...19911722558
promoters in the transfer region of plasmid f. 19853893413
amplification of the respiratory nadh dehydrogenase of escherichia coli by gene cloning.a relatively simple method has been used to clone the gene coding for the respiratory nadh dehydrogenase (nadh-ubiquinone oxidoreductase) of escherichia coli from unfractionated chromosomal dna. the restriction endonucleases ecori, bami and hindiii were used to construct three hybrid plasmid pools from total e. coli dna and the amplifiable plasmids psf2124 and pgm706. three different restriction endonucleases were used to increase the chances of cloning the ndh gene intact. mobilization by the p ...1978365690
construction and some properties of packageable plasmid f.a derivative of plasmid f which is packageable in lambda phage coat was constructed using techniques of in vitro recombination. this plasmid is composed of three dna fragments generated by restriction enzyme ecori: a minif fragment (fragment f5 of f'lac) which is able to replicate autonomously, a dna fragment from staphylococcus plasmid that carries the beta-lactamase gene, and a portion of guaa (b) transducing lambda phage dna carrying lambda cohesive ends (cos site) along with almost all the ...1979286145
indirect sos induction is promoted by ultraviolet light-damaged minif and requires the minif lyna locus.indirect prophage induction is produced by transfer to recipients of u.v.-damaged f plasmid (95 kb). we tested whether the sos signal can be produced by minif, a 9.3 kb restriction fragment, coding for the replication and segregation functions of plasmid f. we used lambda minif, a hybrid phage-plasmid. u.v.-irradiated lambda minif induced prophages phi 80 or lambda and sfia, a chromosomal sos gene, in more than 50% of the infected cells. the maximal inducing dose produced about 0.5 pyrimidine di ...19846096551
location of an f-pilin pool in the inner membrane.polyacrylamide gel analysis of [35s]methionine-labeled membrane preparations from escherichia coli has revealed the presence of five polypeptides present only in the membranes of cells containing the conjugative plasmid f. in addition to the previously reported product of trat, polypeptides migrating with apparent molecular weights of 100,000, 23,500, 12,000, and 7,000 were resolved. membrane preparations from f traj mutants lacked these polypeptides, indicating that all of these proteins are tr ...19816111549
location of f plasmid transfer operon genes trac and traw and identification of the traw product.as part of an analysis of the conjugative transfer genes associated with the expression of f pili by plasmid f, we have investigated the physical location of the trac and traw genes. we found that plasmid clones carrying a 2.95-kilobase ecori-ecorv f transfer operon fragment were able to complement transfer of f lac trac mutants and expressed an approximately 92,000-dalton product that comigrates with trac. we also found that traw-complementing activity was expressed from plasmids carrying a 900 ...19872889720
participation of hup gene product in ori2-dependent replication of fertility plasmid f.the fertility plasmid f'gal was not stably maintained in a hupa-hupb double mutant of escherichia coli. moreover, mini-f plasmids pfzy1, pftc1 and pftc2 were unable to transform the double mutant, though these plasmids efficiently transformed cells harboring a hupa or hupb single mutation. the composite plasmid pfhs1, which consists of the f5 dna fragment of f plasmid and the whole dna of a psc101 derivative that carries a temperature-sensitive mutation for dna replication, was not stably mainta ...19883063607
aspects of plasmid f maintenance in escherichia coli.a major class of replicons in procaryotes is typified by low copy number, nonrandom intracellular distribution, and stable inheritance. included in this class are chromosomes of gram-positive and gram-negative bacteria as well as a number of plasmids from these organisms. replicons in this major class have remarkable structural and functional similarities in the genes that effect and control replication. in the present work a review of plasmid f is presented as a paradigm for many aspects of thi ...19883052759
expression of f transfer functions depends on the escherichia coli integration host factor.we present evidence that the escherichia coli dna binding protein, ihf, plays an important role in conjugal transfer of the plasmid f. our results suggest that ihf exerts this effect by positively effecting transcription of the transfer (tra) operon of the plasmid.19873302598
characterization of the f-plasmid conjugative transfer gene trau.we characterized the trau gene of the escherichia coli k-12 conjugative plasmid f. plasmids carrying segments of the f transfer operon were tested for their capacity to complement f lac trau526. the protein products of trau+ clones were identified, and the nucleotide sequence of trau was determined. trau mapped between traw and trbc. it encodes a 330-amino-acid, mr36,786 polypeptide that is processed. ethanol caused accumulation of a precursor polypeptide; removal of ethanol permitted processing ...19902198250
identification and characterization of the products from the traj and tray genes of plasmid r100.the nucleotide sequence of part of the tra region of r100 including traj and tray was determined, and the products of several tra genes were identified. the nucleotide sequence of traj, encoding a protein of 223 amino acids, showed poor homology with the corresponding segments of other plasmids related to r100, but the deduced amino acid sequences showed low but significant homology. the first four amino acids at the n-terminal region of the traj protein were not essential for positive regulatio ...19882836369
supercoils in prokaryotic dna restrained in vivo.cells of escherichia coli containing the plasmid f were gamma-irradiated with various doses to introduce determined numbers of single-strand breaks in the f dna. the cells were then incubated to permit repair of the breaks while dna gyrase was inhibited with coumermycin to limit restoration of any relaxed supercoil. repaired, covalently continuous f dna was isolated and its superhelical density was measured by two different methods. both indicated that a major part (50-60%) of the negative super ...19806246488
mini-f e protein: the carboxy-terminal end is essential for e gene repression and mini-f copy number control.mini-f is a segment of the conjugative plasmid f consisting of two origins of replication flanked by regulatory regions, which ensure a normal control of replication and partitioning. adjacent to the ori-2 origin is a complex coding region that consists of the e gene overlapped by three open reading frames with the coding potential for 9000 mr polypeptides here designated 9 kd-1, 9 kd-2 and 9 kd-3. in this paper, we show that open reading frame 9 kd-3 is preceded by active promoter and shine-dal ...19863018261
phage folac: an folac plasmid-dependent bacteriophage.by enriching sewage with escherichia coli or salmonella typhimurium strains harbouring the plasmid edp208, a constitutive pilus-producing derivative of plasmid f olac, a phage was isolated which plated on these two organisms but not on isogenic strains without the plasmid. the phage was named f olac; it has a hexagonal outline with a diameter of 28 nm, contained rna, was resistant to chloroform, and probably adsorbed preferentially to the sides of edp208 pili very near the tip. phage multiplicat ...19816121841
[isolation and properties of recombinant dna from a plasmid and filamentous phage].in the course of studying extrachromosomal dna with composite replicons, a hybrid has been constructed by the in vitro recombination of the filamentous phage m13mp2 dna (rf) and plasmid pur222 (apr). both parental dnas contain a fragment of lac-operon (ca. 800 bp), which includes the distal end of laci gene, lacpo segments, and the lacz gene proximal region coding for 145 n-terminal amino acid residues of beta-galactosidase and thus providing for alpha-complementation, the effect being cancelled ...19863004511
two functions of the e protein are key elements in the plasmid f replication control system.by using a plasmid carrying a translational fusion between the e gene of the incfi plasmid f and the lacz gene, we located the operator of the autogenously regulated e gene to an inverted repeat overlapping the e-gene promoter and showing perfect homology to part of the sequence found in all the direct repeats of two regions exerting an inhibitory effect on f replication, incb and incc. excess e protein provided in trans to an f plasmid increased the replication frequency of the f plasmid. this ...19852999077
tn2301, a transposon construct carrying the entire transfer region of the f plasmid.the largest r . bamhi fragment of the plasmid f, which carries the entire f conjugation system, has been cloned into the single r . bamhi site of the ampicillin (ap) resistance transposon tn1. pds1106 (cole1 mob::tn1) was the vector plasmid, and the resultant conjugative plasmid, ped830, was characterized both genetically and by restriction enzyme analysis. the transposon construct, denoted tn2301, was transposable at frequencies similar to tn1 to small nonconjugative plasmids or to the escheric ...19806251027
stability of a plasmid f trim in populations of a recombination-deficient strain of escherichia coli in continuous culture.populations of a reca derivative of escherichia coli ab1157 containing the plasmid f trim were grown in carbon-limited continuous culture at dilution rates of 0.1 h-1 to 0.4 h-1. the plasmid was lost after a lag, except in fermenter-experienced populations when it was retained. these results can be explained in terms of non-specific competition.19846380407
recombinational instability of f' plasmids in escherichia coli k-12: localization of fre-sites.the f' plasmids orf-1 (pure+ tsxs proc+ lac+) and f'14 (arge+ metb+ ilv+) contain active regions of recombination, fre i and fre ii correspondingly. the plasmid orf-1 is stable in recf- cells (i.e., with the recbc pathway of recombination) and decays in rec+ cells (recbcf pathway) giving two types of product: f+ and plasmid pck-1 (tsxs proc+ lac+) containing part of the initial dna. they are extremely instable in the presence of the recf pathway, (recbc- sbcb-), yielding f+ and plasmid pck-2 (pr ...19816276675
polar mobilization of the escherichia coli chromosome by the cole1 transfer origin.mobilization of the plasmid cole1 from cells containing a conjugative plasmid (such as f) requires the synthesis of cole1 mob proteins, and the presence, in cis, of bom (basis of mobility), a region of cole1 containing the origin of transfer (orit). the process of cole1 transfer is thought to resemble that of the conjugative plasmid f, although the plasmids share little sequence homology. in f, conjugation is preceded by a strand-specific nicking event at orit. the nicked strand is then conducte ...19863018432
[participation of plasmid f in the replication of the chromosome of an hfr strain of escherichia coli k-12].in the region of plasmid f dna with coordinates 52,2-55,8 kb, the chr ("chromosome replication") locus has been revealed. a failure in the functioning of this locus in the integrated plasmid, which leads to a temperature-sensitive disturbance in chromosome replication of the hfr strain and to the changes in its sensitivity to some membranotropic agents. integration of an f segment containing the chr+ allele into the chromosome of an f-like derivative of such hfr strain (retaining a mutant part o ...19836343185
a versatile low-copy-number cloning vector derived from plasmid f.we have constructed a cloning vector based on plasmid mini-f for use in escherichia coli. plasmid pzc320 consists of the ori-2 replication unit of f that confers very low copy number (lcn), and includes the sop partition functions to insure stable plasmid maintenance in the absence of selection. a multiple cloning site (mcs) containing 16 unique restriction sites is located within the 5' end of the lacz alpha gene. expression of lacz alpha is under the control of the wild-type lactose operator/p ...19957590321
consequences of interaction between f plasmid and a drug-resistance plasmid belonging to incompatibility group f1.two plasmids, plk1 and plk2, were derived from pip218, an in vivo recombinant of plasmid f and the drug-resistance plasmid pip176 (cmr, smr, sur, tcr). of these two plasmids, plk1 is 70 mdaltons and carries the tc-resistance determinant in a 7-mdalton transposition element; plk2 is 125 mdaltons and carries cm-, sm- and su-resistance determinants. the plasmid plk1 resulted as a tc-resistance segregaant of pip218 during its coexistence with another plasmid, co1e1-arac101, and plk2 (125 mdaltons) a ...19806250688
recombinant dna risk assessment studies in humans: efficacy of poorly mobilizable plasmids in biologic containment.recombinant dna risk assessment studies quantitated the mobilizability of "safe" plasmid pbr325, in comparison with readily mobilizable plasmid pjbk5 (chloramphenicol and tetracycline resistant). of 15 volunteers who became colonized after ingestion of 5 x 10(10) escherichia coli hs-4, a normal human flora strain containing pjbk5 and daily oral tetracycline, nine manifested transfer of pjbk5 to normal flora by means of triparental mating. in contrast, none of 12 other volunteers cocolonized with ...19836355313
identification of the minimal essential region for the replication origin of minif plasmid.the minimal replication origin of minif plasmid was found to lie within a region of 217 bp in length. this region contains an at cluster and the four 19 bp direct repeats responsible for incompatibility, termed incb. its location coincides with hat of ori2 of plasmid f, previously inferred to be the replication starting point by electron microscopic analysis (eichenlaub et al. 1981).19846593565
mutagenic and recombinagenic effects of diethylstilbestrol quinone.estrogens are believed to be major contributors to many cancers of the human female genital tract, but the mechanism of their carcinogenic action is not well-understood. while a tumor-promoting role for estrogens is well-supported, whether they also act as tumor initiators has remained controversial. here, we have sought to examine the mutagenic potential of diethylstilbestrol, a synthetic estrogen that is a powerful carcinogen in hamsters, and is suspected to be a human carcinogen. phage m13 si ...19937690889
promiscuous dna transfer system of agrobacterium tumefaciens: role of the virb operon in sex pilus assembly and synthesis.conjugative transfer of dna that occurs between bacteria also operates between bacteria and higher organisms. the transfer of dna between gram-negative bacteria requires initial contact by a sex pilus followed by dna traversing four membranes (donor plus recipient) using a transmembrane pore. accumulating evidence suggests that transfer of the t-dna from agrobacterium tumefaciens to plants may also occur via a conjugative mechanism. the virb operon of the ti plasmid exhibits close homologies to ...19947914664
cyclic amp and its receptor protein are required for expression of transfer genes of conjugative plasmid f in escherichia coli.a number of cya and crp mutants of escherichia coli hfrh were analyzed for several tra functions of the f plasmid. the mutants were observed to be deficient in conjugal donor ability, absorption of phages ms2 and q beta and surface exclusion. these defects were suppressed in cya mutants grown with camp supplementation. a camp concentration of 3 x 10(-4) m produced maximal suppression of donor ability defect in a cya strain. camp did not suppress the tra- phenotype of crp mutants. latent periods ...19836304473
overproduction in escherichia coli k-12 and purification of the traj protein encoded by the conjugative plasmid f.the traj protein encoded by the conjugative plasmid f has been designated an escherichia coli k-12 envelope protein that participates in a mechanism of gene regulation. we have purified the traj protein, using an flac::lambda traj lysogen that overproduces the protein after heat induction of the prophage. sufficient traj protein was synthesized within 40 min after induction to follow the purification by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. our purification exploited the obs ...19846327690
genetic analysis of escherichia coli k-12 chromosomal mutants defective in expression of f-plasmid functions: identification of genes cpxa and cpxb.two temperature-sensitive, chromosomal mutants of escherichia coli were selected for their inability to express deoxyribonucleic acid donor activity and other activities associated with the conjugative plasmid f. these mutants were also auxotrophic for isoleucine and valine at 41 degrees c. each mutant strain contained two altered genes: cpxa, located at 88 min on the e. coli k-12 genetic map, and cpxb, located at 41 min. mutations in both genes were required for maximal expression of mutant phe ...19806998969
regulated expression of a gene important for replication of plasmid f in e. coli.fusions between the gene encoding the e protein of the incfi plasmid f and the lac genes were constructed. analysis of the expression of beta-galactosidase from these fusions shows that the promoter for the e protein gene is located between the incb region and the structural gene for the e protein. near this promoter is a regulatory site on which a negative control element acts. most likely the e protein itself acts as a repressor of e gene expression and thus autoregulates its own expression. n ...19846325163
purification and biochemical characterization of trwc, the helicase involved in plasmid r388 conjugal dna transfer.trwc is an essential protein in conjugative dna transfer of the broad-host-range plasmid r388. trwc was purified in two chromatographic steps from trwc-overproducing bacteria. the purification procedure resulted in > 90% pure trwc protein, which was free of contaminating nuclease activities. trwc behaved as a dimer in gel-filtration chromatography in the presence of 550 mm nacl, and had a pi of 10.1. the purified protein showed in-vitro ssdna-dependent nucleoside-5'-triphosphatase and dna helica ...19948001558
plasmids isolated from marine sediment microbial communities contain replication and incompatibility regions unrelated to those of known plasmid groups.two hundred ninety-seven bacteria carrying plasmids that range in size from 5 to 250 kb were identified from more than 1,000 aerobic heterotrophic bacteria isolated from coastal california marine sediments. while some isolates contained numerous (three to five) small (5- to 10-kb) plasmids, the majority of the natural isolates typically contained one large (40- to 100-kb) plasmid. by the method of plasmid isolation used in this study, the frequency of plasmid incidence ranged from 24 to 28% depe ...19979055407
escherichia coli coiv plasmid prk100: genetic organization, stability and conjugal transfer.uropathogenic escherichia coli strains express chromosomal and plasmid-encoded virulence-associated factors such as specific adhesins, toxins and iron-uptake systems. a coiv plasmid (prk100) of a uropathogenic strain and its host ks533 were studied. the host strain encodes the k1 capsule, and p and s fimbriae, but neither haemolysin nor the cytotoxic-necrotic factor cnf1, indicating that this strain does not harbour a larger pathogenicity island. a restriction map of prk100 was constructed on th ...19989493372
a proposed system for nomenclature for incompatibility genes of the escherichia coli sex factor, plasmid f. 19806765585
random initiation of replication of plasmids p1 and f (oris) when integrated into the escherichia coli chromosome.we have constructed intp1 and intfs strains of escherichia coli in which the basic replicons of either plasmid p1 or plasmid f (oris) were integrated into an inactivated oric, such that chromosome replication is controlled by the integrated plasmid replicon. in this study, we have further analysed these strains, and density-shift experiments revealed that chromosome replication occurred randomly during the cell cycle. flow-cytometry analyses of exponentially growing populations supported this co ...19968809755
subdomain organization and catalytic residues of the f factor trai relaxase domain.trai from conjugative plasmid f factor is both a "relaxase" that sequence-specifically binds and cleaves single-stranded dna (ssdna) and a helicase that unwinds the plasmid during transfer. using limited proteolysis of a trai fragment, we generated a 36-kda fragment (trai36) retaining trai ssdna binding specificity and relaxase activity but lacking the ssdna-dependent atpase activity of the helicase. further proteolytic digestion of trai36 generates stable n-terminal 26-kda (trai26) and c-termin ...200312637015
dna recognition by f factor trai36: highly sequence-specific binding of single-stranded dna.the trai protein has two essential roles in transfer of conjugative plasmid f factor. as part of a complex of dna-binding proteins, trai introduces a site- and strand-specific nick at the plasmid origin of transfer (orit), cutting the dna strand that is transferred to the recipient cell. trai also acts as a helicase, presumably unwinding the plasmid strands prior to transfer. as an essential feature of its nicking activity, trai is capable of binding and cleaving single-stranded dna oligonucleot ...200111560509
the f plasmid ccdb protein induces efficient atp-dependent dna cleavage by gyrase.dna topoisomerases perform essential roles in dna replication, gene transcription, and chromosome segregation. recently, we identified a new type of topoisomerase ii poison: the ccdb protein of plasmid f. when its action is not prevented by ccda protein, the ccdb protein is a potent cytotoxin. in this paper, using purified ccdb, ccda and gyrase, we show that ccdb protein efficiently traps gyrase in a cleavable complex. the ccda protein not only prevents the gyrase poisoning activity of ccdb but ...19938254658
the product of the virb4 gene of agrobacterium tumefaciens promotes accumulation of virb3 protein.the process of t-dna transfer from agrobacterium tumefaciens to plant cells is thought to involve passage of a dna-protein complex through a specialized structure in the bacterial membrane. the virb operon of a. tumefaciens encodes 11 proteins, of which 9 are known to be located in the membranes and 10 have been shown to be essential for virulence. sequence comparisons between proteins encoded by the virb operon and those encoded by operons from conjugative plasmids indicated that virb proteins ...19948071199
a hot spot in plasmid f for site-specific recombination mediated by tn21 integron integrase.integron in2 integrase (inti1)-mediated site-specific recombination between two primary sites occurs at a high frequency, while that between a primary and a secondary site occurs at frequencies around 10,000 times lower. secondary sites consist of a pentanucleotide with only two fully conserved residues (gwtmw). the analysis of inti1-mediated recombinants in the plasmid pox38 revealed the existence in this plasmid of a site used at a frequency intermediate between those of primary and secondary ...19979209065
escherichia coli strains in which chromosome replication is controlled by a p1 or f replicon integrated into oric.we report the construction of intp1 and intfs strains, in which the basic replicon from either plasmid p1 or plasmid f (oris) has been integrated in both orientations into the origin of replication, oric, of the escherichia coli chromosome. in these strains, oric is no longer functional and chromosome-replication is instead controlled by the integrated plasmid replicon. the strains were viable, showing that the deviation from normal chromosome-replication control was not large enough to prohibit ...19968809754
genetic organization of the conjugal dna processing region of the incw plasmid r388.the region of the incw plasmid r388 involved in conjugal dna metabolism and mobilization (mobw) has been analyzed by tn5tac1 insertion mutagenesis, genetic complementation and dna sequencing. three genes (trwa, trwb and trwc) were mapped within mobw. they are transcribed from the same strand and away from orit. the predicted products of trwa, trwb and trwc are proteins of 121, 507 and 966 amino acids, respectively. the three proteins were visualized in a minicell expression system, showing appar ...19948289274
a putative lichenysin a synthetase operon in bacillus licheniformis: initial characterization.certain bacillus licheniformis strains isolated from oil wells have been shown to produce a very effective biosurfactant, lichenysin a, which is structurally similar to another less active lipopeptide, surfactin. surfactin, like many small peptides in prokaryotes and lower eukaryotes, is synthesized non-ribosomally by multi-enzyme peptide synthetase complex. analysis of several peptide synthetases of bacterial and fungal origin has revealed a high degree of sequence conservation. two 35-mer olig ...19989765590
bacteriophage mu repressor as a target for the escherichia coli atp-dependent clp protease.bacteriophage mu repressor, which is stable in its wildtype form, can mutate to become sensitive to its escherichia coli host atp-dependent clpxp protease. we further investigated the determinants of the mutant repressor's sensitivity to clp. we show the crucial importance of a c-terminal, seven amino acid long sequence in which a single change is sufficient to decrease the rate of degradation of the protein. the sequence was fused at the c-terminal end of the ccdb and ccda proteins encoded by p ...19968617219
isolation and identification of fxsa, an escherichia coli gene that can suppress f exclusion of bacteriophage t7.a selection for mutants of escherichia coli that survive coexpression of bacteriophage t7 gene 10 and plasmid f pifa has allowed the identification of a newly defined genetic locus, fxsa. fxsa is located at 94.1 min on the e. coli chromosome; the gene is monocistronic and non-essential for growth. overexpression of fxsa is necessary for resistance to the toxicity of t7 gene 10 in the presence of pifa; the original mutant strain contains a promoter-up mutation, changing a g residue to the "invari ...199910497016
the plasmid f ompp protease, a homologue of ompt, as a potential obstacle to e. coli-based protein production.ompt, an outer membrane-localized protease of escherichia coli, cleaves a number of exogenous and endogenous proteins during their purification. secy, an endogenous membrane protein, is a target of this artificial proteolysis in vitro. here we report that secy cleavage occurs even in cell extracts from ompt-disrupted cells, if they carry an f plasmid derivative. a gene, ompp, on the f plasmid was shown to be responsible for this proteolysis. these results indicate that the absence of an f-like p ...199910561486
virus-plasmid interactions: mutants of bacteriophage t3 that abortively infect plasmid f-containing (f+) strains of escherichia coli.bacteriophage t7 and many closely related phages abortively infect plasmid f-containing (f+) strains of escherichia coli. however phage t3, which is also closely related to t7, grows normally in f+ hosts. mutants of phage t3 that, like t7, are subject to f-mediated restriction have been isolated. these t3 mutants lack or are defective in one or both of two genes that are nonessential for phage growth in f-, wild-type strains. our results show that the products of phage t3 gene 1.1 or 1.2, or bot ...19846324192
crystallization and preliminary x-ray characterization of the relaxase domain of f factor trai.conjugative plasmids are capable of transferring a copy of themselves in single-stranded form from donor to recipient bacteria. prior to transfer, one plasmid strand must be cleaved in a sequence-specific manner by a relaxase or mobilization protein. trai is the relaxase for the conjugative plasmid f factor. a 36 kda n-terminal fragment of trai possesses the single-stranded dna-binding and cleavage activity of the protein. crystals of the 36 kda trai fragment in native and selenomethionine-label ...200312876370
non-cytotoxic variants of the kid protein that retain their auto-regulatory activity.kid and kis are, respectively, the toxin and antitoxin encoded by the pard operon of plasmid r1. the recently solved crystal structure of kid has revealed that this protein closely resembles the ccdb toxin of plasmid f. in ccdb, the residues involved in toxicity are located at the carboxy-terminal end of the protein. however, an analogous information on the kid toxin was not available. here, we have characterized a collection of non-toxic mutants of the kid protein and identified the residues th ...200312932738
plasmids of the same inc groups in enterobacteria before and after the medical use of antibiotics.conjugative plasmids were common in enterobacteria isolated before the medical use of antibiotics. plasmid f of escherichia coli k-12 was one example and we identified others in over 20% of a collection of strains isolated between 1917 and 1954, the murray collection. in the past 25 years, conjugative plasmids encoding antibiotic resistances have become common in bacteria of the same genera as those of the murray collection--salmonella, shigella, klebsiella, proteus, escherichia. the present stu ...19836316165
lon-dependent proteolysis of ccda is the key control for activation of ccdb in plasmid-free segregant bacteria.the ccd locus contributes to the stability of plasmid f by post-segregational killing of plasmid-free bacteria. the ccdb gene product is a potent cell-killing protein and its activity is negatively regulated by the ccda protein. in this paper, we show that the ccda protein is unstable and that the degradation of ccda is dependent on the lon protease. differences in the stability of the killer ccdb protein and its antidote ccda are the key to post-segregational killing. because the half-life of a ...19948022284
crystal structure of a prokaryotic replication initiator protein bound to dna at 2.6 a resolution.the initiator protein (repe) of f factor, a plasmid involved in sexual conjugation in escherichia coli, has dual functions during the initiation of dna replication which are determined by whether it exists as a dimer or as a monomer. a repe monomer functions as a replication initiator, but a repe dimer functions as an autogenous repressor. we have solved the crystal structure of the repe monomer bound to an iteron dna sequence of the replication origin of plasmid f. the repe monomer consists of ...199910469640
[the study of heterogeneity of plasmid-bearing and plasmid-f ree populations of bacillus subtilis under different environmental conditions].the population heterogeneity of recombinant and plasmid-free bacillus subtilis strains introduced into aquatic microcosms was studied. after introduction, the population of the plasmid-free strain b. subtilis 2335 in microcosms has long been represented by both vegetative cells and spores, whereas, already ten days after introduction, the population of the recombinant strain b. subtilis 2335/105 (km[symbol: see text]nf+) was represented only by spores. the number of plasmid copies in the spore i ...200010776630
inter- and intramolecular determinants of the specificity of single-stranded dna binding and cleavage by the f factor relaxase.the trai protein of conjugative plasmid f factor binds and cleaves a single-stranded region of the plasmid prior to transfer to a recipient. trai36, an n-terminal trai fragment, binds ssdna with a subnanomolar k(d) and remarkable sequence specificity. the structure of the trai36 y16f variant bound to ssdna reveals specificity determinants, including a ssdna intramolecular 3 base interaction and two pockets within the protein's binding cleft that accommodate bases in a knob-into-hole fashion. mut ...200516216584
plasmid distribution in european diaporthe helianthi isolates.diaporthe helianthi is the causal agent of sunflower stem canker, a serious pathogen of sunflower in europe, which has been sporadically recorded in italy. a collection of 26 diaporthe helianthi isolates deriving from different geographic origins was analysed in order to determine the presence of extra-chromosomal genetic determinants and their molecular diversity. extra-chromosomal bands in total genomic dnas were identified in every french and the yugoslavian isolate and in only one italian is ...200515983747
incp plasmids are most effective in mediating conjugation between escherichia coli and streptomycetes.mobilizable shuttle plasmids containing the origin of transfer (orit) region of plasmid f (incfi), colib-p9 (inci1), and rp4/rp1 (incpalpha) were constructed to test the ability of the cognate conjugation system to mediate gene transfer from escherichia coli to streptomyces. the conjugative system of the incpalpha plasmids was shown to be most effective in conjugative transfer, giving peak values of (2.7 +/- 0.2) x 10(-2) s. lividans tk24 exconjugants per recipient cell. to assess whether the ma ...200616808239
the highly conserved tldd and tlde proteins of escherichia coli are involved in microcin b17 processing and in ccda degradation.microcin b17 (mccb17) is a peptide antibiotic produced by escherichia coli strains carrying the pmccb17 plasmid. mccb17 is synthesized as a precursor containing an amino-terminal leader peptide that is cleaved during maturation. maturation requires the product of the chromosomal tlde (pmba) gene. mature microcin is exported across the cytoplasmic membrane by a dedicated abc transporter. in sensitive cells, mccb17 targets the essential topoisomerase ii dna gyrase. independently, tlde as well as t ...200212029038
crystallization of the c-terminal domain of the addiction antidote ccda in complex with its toxin ccdb.ccda and ccdb are the antidote and toxin of the ccd addiction module of escherichia coli plasmid f. the ccda c-terminal domain (ccdac36; 36 amino acids) was crystallized in complex with ccdb (dimer of 2 x 101 amino acids) in three different crystal forms, two of which diffract to high resolution. form ii belongs to space group p2(1)2(1)2(1), with unit-cell parameters a = 37.6, b = 60.5, c = 83.8 a and diffracts to 1.8 a resolution. form iii belongs to space group p2(1), with unit-cell parameters ...200516511204
effect of extremely low frequency magnetic field exposure on dna transposition in relation to frequency, wave shape and exposure time.purpose: to examine the effect of extremely low frequency magnetic field (elf-mf) exposure on transposon (tn) mobility in relation to the exposure time, the frequency and the wave shape of the field applied. materials and methods: two escherichia coli model systems were used: (1) cells unable to express β-galactosidase (lacz(-)), containing a mini-transposon tn10 element able to give ability to express β-galactosidase (lacz(+)) upon its transposition; therefore in these cells transposition activ ...201121504343
hydrodynamic plasmid dna gene therapy model in liver transplantation.there is great interest in the field of transplantation to genetically modify grafts to decrease preservation injury or allograft rejection. although adenoviral gene transfer has been effective in experimental liver transplantation, viral toxicity and safety concerns limit potential use in clinical trials. therefore, the purpose of this study was to develop a model of nonviral gene transfer in the liver transplant setting, allowing for efficient transgene expression.200616926028
polymerization of sopa partition atpase: regulation by dna binding and sopb.in bacteria, mitotic stability of plasmids and many chromosomes depends on replicon-specific systems which comprise a centromere, a centromere-binding protein and an atpase. dynamic self-assembly of the atpase appears to enable active partition of replicon copies into cell-halves, but for most atpases (the walker-box type) the mechanism is unknown. also unknown is how the host cell contributes to partition. we have examined the effects of non-sequence-specific dna on in vitro self-assembly of th ...200717166176
the incc korb region of rk2 repositions a mini-rk2 replicon in escherichia coli.analysis by fluorescence microscopy has established that plasmid rk2 in escherichia coli and other gram-negative bacteria is present as discrete clusters that are located inside the nucleoid at the mid- or quarter-cell positions. a mini-rk2 replicon containing an array of teto repeats was visualized in e. coli cells that express a tetr-eyfp fusion protein. unlike intact rk2, the rk2 mini-replicon (pcv1) was localized as a cluster at the cell poles outside of the nucleoid. insertion of the o(b1)i ...200717521722
insight into centromere-binding properties of parb proteins: a secondary binding motif is essential for bacterial genome maintenance.parb proteins are one of the three essential components of partition systems that actively segregate bacterial chromosomes and plasmids. in binding to centromere sequences, parb assembles as nucleoprotein structures called partition complexes. these assemblies are the substrates for the partitioning process that ensures dna molecules are segregated to both sides of the cell. we recently identified the sopc centromere nucleotides required for binding to the parb homologue of plasmid f, sopb. this ...201323345617
f plasmid partition depends on interaction of sopa with non-specific dna.bacterial atpases belonging to the para family assure partition of their replicons by forming dynamic assemblies which move replicon copies into the new cell-halves. the mechanism underlying partition is not understood for the walker-box atpase class, which includes most plasmid and all chromosomal paras. the atpases studied both polymerize and interact with non-specific dna in an atp-dependent manner. previous work showed that in vitro, polymerization of one such atpase, sopa of plasmid f, is i ...200818826408
replication of a unit-copy plasmid f in the bacterial cell cycle: a replication rate function analysis.for stability, the replication of unit-copy plasmids ought to occur by a highly controlled process. we have characterized the replication dynamics of a unit-copy plasmid f by a replication rate function defined as the probability per unit age interval of the cell cycle that a plasmid will initiate replication. analysis of baby-machine data [j. bacteriol. 170 (1988) 1380; j. bacteriol. 179 (1997) 1393] by stochastics that make no detailed reference to underlying mechanism revealed that this rate ...200415212889
a plasmid rk2-based broad-host-range cloning vector useful for transfer of metagenomic libraries to a variety of bacterial species.the majority of microorganisms in natural environments are difficult to cultivate, but their genes can be studied via metagenome libraries. to enhance the chances that these genes become expressed we here report the construction of a broad-host-range plasmid vector (prs44) for fosmid and bacterial artificial chromosome (bac) cloning. prs44 can be efficiently transferred to numerous hosts by conjugation. it replicates in such hosts via the plasmid rk2 origin of replication, while in escherichia c ...200919459950
interaction of the icia protein with at-rich regions in plasmid replication origins.a set of at-rich repeats is a common motif in prokaryotic replication origins. we have screened for proteins binding to the at-rich repeat region in plasmids f, r1 and psc101 using an electrophoretic mobility shift assay with pcr-amplified dna fragments from the origins. the icia protein, which is known to bind to the at-rich repeat region in the escherichia coli origin of chromosome replication, oric, was found to bind to the corresponding region from plasmids f (oris) and r1, but not to psc101 ...19968657567
energetics of the sequence-specific binding of single-stranded dna by the f factor relaxase domain.transfer of conjugative plasmids between bacteria requires the activity of relaxases or mobilization proteins. these proteins nick the plasmid in a site- and strand-specific manner prior to transfer of the cut strand from donor to recipient. trai36, the relaxase domain of trai from plasmid f factor, binds a single-stranded dna (ssdna) oligonucleotide containing an f factor sequence with high affinity and sequence specificity. to better understand the energetics of this interaction, we examined t ...200415123728
the activity of a single-stranded promoter of plasmid colib-p9 depends on its secondary structure.the leading region of the conjugal bacterial plasmid colib-p9 contains three dispersed repeats of a 328 bp sequence homologous to frpo, a sequence from plasmid f that acts as a promoter in single-stranded dna. one of these sequences, ssi3, inactive in the double-stranded form, promoted in vitro transcription exclusively from the single strand that is transferred during conjugation. promoter activity was dependent on the presence of rna polymerase holoenzyme containing sigma 70. transcription ini ...200415228523
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