Nucleotide sequence of the filamentous bacteriophage M13 DNA genome: comparison with phage fd

Wezenbeek, Peter M.G.F. van; Hulsebos, Theo J.M.; Schoenmakers, John G.G.

doi:10.1016/0378-1119(80)90093-1

Cited by 404 publications

(166 citation statements)

References 47 publications

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“…A similar conclusion can be drawn from a pausing site analysis of the gene 1 region (position 3406-3200) ofM13nipX (+) DNA. A very stable secondary structure had been proposed earlier for this region a t position 3341 [22], which is also predicted by our computer analysis. With the gene 1 primer which hybridizes to position 3406 -3420 we obscrvc two pronounced pausing sites at positions 3344 and 3343, one and t w o nucleotides apart from the hairpin stem (Fig.…”

Section: Muppftig Of'ssecondury Structuressupporting

confidence: 90%

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Analysis of secondary structures in M13mp8 (+) single‐stranded DNA by the pausing of DNA polymerase α

Reckmann

Grosse

Urbanke

et al. 1985

European Journal of Biochemistry

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The pausing of DNA replication has been used as a tool for analyzing secondary structures in a single-stranded DNA. M13mp8 (+) single-stranded DNA was replicated in vitro by the DNA polymerase x from calf thymus. The positions of pausing were determined from DNA sequencing gels. All experimentally observed pausing sites could be correlated with computer-predicted secondary structures of the MI3 single-stranded DNA. In the computer calculations of the secondary structures, long-range base-pairing, Ci. T mispairs and loop-out of bases were allowed. By using six different primers, the pausing site pattern and the corresponding secondary structure map o f a region comprising 3400 nucleotides of the M I 3 genome has been established. Our experiments indicate that the M I 3 TINA is highly structured. Most of the stable structures are clustered around the origin ofreplication. With fragments of the M13 DNA, we show that long-range base-pairing exists in the M13 single-strandcd gcnomc and we present evidence for tertiary structure interactions. Furthermore we observe structures that form newly during the course of replication. The Eschcrichiu coli single-stranded DNA-binding protein facilitates replication through the barriers.The in virro replication of natural single-stranded DNA has proved to be an effective tool for studying the replicative properties of DNA polymerases. In these experiments characteristic stops of replications have been noted at defined positions on the singlc-stranded D N A templates. Such pausing has been described for many polymerases as, for example, DNA polymerase I1 [l] and DNA polymerasc I11 from Estherichia coli [2], bacteriophage T4 DNA polymerase [3, 41, vaccinia virus polymerase [ 5 ] , DNA polymerase M from D~o~ophilu melunoguster [6, 71 and from CV-1 monkey cells [8]. The pausing sites have been analyzed to one-nucleotide resolution by means of DNA sequencing gels [2, 7, 81. It has been shown in several systems that stable secondary structures of the single-stranded D N A template can lead to a pausing of DNA polymerases [l, 3, 61. However, only part of the experimentally observed pausing sites could be correlated with secondary structures. Hence, pausing has been attributed also to primary sequence effects [2, 7, 81. Studies on the nature of the pausing sites have been hampered by lack of knowledge about the secondary structures in large single-stranded DNAs. Since experimental data are not available, one has to rely on computer calculations. In our computer calculations we have chosen to consider incorrect base pairs, loop-out of single bases and longrange interactions. Using the in uitro replication of M 13 singlestranded DNA by the 9s DNA polymerase 3 from calf thymus, we have analyzed the pausing sites over a range of about 1400

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Section: Muppftig Of'ssecondury Structuressupporting

confidence: 90%

“…7 ) . The corresponding computer-predicted hairpin structures, which largely verify the earlier proposals for secondary structures in this region [22], are shown in Fig. 8.…”

Section: G -Csupporting

confidence: 79%

Analysis of secondary structures in M13mp8 (+) single‐stranded DNA by the pausing of DNA polymerase α

Reckmann

Grosse

Urbanke

et al. 1985

European Journal of Biochemistry

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“…Upstream, the remainder of the 870 bp small fragment is unrelated 6-lactamase coding sequence which does not contain a promoter, nor is it transcribed by readthrough from an upstream phage promoter (46,47). These observations strongly suggest that each point mutation in 5PR-01, -04 and -05 is responsible for the increase in expression.…”

Section: Vector Constructionmentioning

confidence: 99%

Increased expression of a cloned gene by local mutagenesis of Its promoter and ribosome binding site

Warburton¹,

Boseley²,

Porter³

1983

Nucl Acids Res

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A strategy for local mutagenesis of DNA has been developed. The lac promoter in phage M13mp9 was replaced with the E.coli trp promoter. A restriction fragment bearing only the trp promoter region was mutagenized with nitrous acid, religated to the unmutagenized vector and transfected into E.coli. Several clones which give darker blue plaques on indicator media, suggesting increased a-galactosidase synthesis, were selected for DNA sequencing. One clone has a G--A transition on the 3' side of the 'Pribnow box' which results in a constitutive promoter. Two clones have different point mutations (C--T and T--C) between the Shine-Dalgarno sequence and initiation codon which raise expression of a-galactosidase two-fold.A secondary structure model suggests that the latter two mutations could exert their effect by destabilizing base-pairing of the lac Z coding region with the ribosome binding site (RBS), thereby allowing easier access to ribosomes. Support for the model comes from the finding that neither of the RBS mutations increase expression of a different downstream gene which forms no obvious secondary structure with the RBS region, whether or not the mutations are present.These results strengthen the hypothesis that secondary structure masking is a major determinant of RBS strength.

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“…Calcium chloride-treated LA6 cells were then transfected with DNA from the ligation reaction and plated for plaque formation. After phage DNAs from 100 small plaques were analyzed by a "rapid gel method" (11), 13 20 min at 370C, the cultures were harvested and cell lysates were analyzed by sucrose density gradients as described (4). The direction of sedimentation is from left to right (horizontal arrow in a).…”

mentioning

confidence: 99%

Identification of ColE1 DNA sequences that direct single strand-to-double strand conversion by a phi X174 type mechanism.

Nomura¹,

Low²,

Ray³

1982

Proc. Natl. Acad. Sci. U.S.A.

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A DNA single-strand initiation sequence, named rriA (called rm-i previously), was detected in the origin region (Hae II fragment E) of the ColEl plasmid [Nomura, N. & Although the mechanism of leading strand (L-strand) initiation in ColEl replication has been studied extensively (1,2), that ofthe lagging strand (H-strand) is still unclear. Because L-strand initiation creates a nascent 6S L-strand fragment to form, a D loop (3), initiation of H-strand synthesis might take place on the displaced, parental single-stranded L-strand DNA (ss DNA) template. This mechanism suggests that genetic information necessary for initiation of H-strand replication resides in this L-strand sequence and may direct initiation in a manner similar to that used by one of the ss DNA phage of Escherichia coli. To explore this possibility, the origin region fragment (Hae II fragment E) of ColEl DNA has been cloned into the filamentous DNA phage M13, and a site capable of promoting DNA strand initiation on a ss DNA template has been identified on the L-strand of this fragment (4). The site, named rriA, directs conversion of chimeric phage ss DNA to parental replicative form (RF) in the presence of rifampicin, an inhibitor ofthe RNA polymerase-dependent mechanism normally used by M13. The dnaB and dnaG gene products were found to be required for function of rriA in vivo (4).Two mechanisms of dnaG-dependent primer synthesis have been identified by in vitro DNA replication studies of small ss DNA phage. In G4 complementary strand synthesis, primase (dnaG protein) synthesizes a unique RNA primer (5). In 4X174

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Nucleotide sequence of the filamentous bacteriophage M13 DNA genome: comparison with phage fd

Cited by 404 publications

References 47 publications

Analysis of secondary structures in M13mp8 (+) single‐stranded DNA by the pausing of DNA polymerase α

Analysis of secondary structures in M13mp8 (+) single‐stranded DNA by the pausing of DNA polymerase α

Increased expression of a cloned gene by local mutagenesis of Its promoter and ribosome binding site

Identification of ColE1 DNA sequences that direct single strand-to-double strand conversion by a phi X174 type mechanism.

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