Bacteriophage PM2 DNA Research Articles

The binding of echinomycin to deoxyribonucleic acid.

Echinomycin is a peptide antibiotic which binds strongly to double-helical DNA up to a limit of approximately one molecule per five base-pairs. There is no detectable interaction with rRNA and only extremely feeble non-specific interaction with poly(rA)-poly(rU). Heat denaturation of DNA greatly decreases the binding, and similarly limited interaction is observed with naturally occurring single-stranded DNA. Association constants for binding to nine double-helical DNA species from different sources are presented; they vary by a factor of approximately 10, but are not simply related to the gross base composition. The interaction with DNA is ionic-strength-dependent, the binding constant falling by a factor of 4 when the ionic strength is raised from 0.01 to 0.10mol/litre. From the effect of temperature on the association constant for calf thymus DNA, the enthalpy of interaction is calculated to be about -13kJ/mol (-3kcal/mol). Binding of echinomycin persists in CsCl gradients and the buoyant density of nicked bacteriophage PM2 DNA is decreased by 25 mg/ml. Echinomycin interacts strongly with certain synthetic poly-deoxynucleotides, the binding constant decreasing in the order poly(dG)-poly(dC) greater than poly(dG-dC) greater than poly(dA-dT). For the latter two polymers the number of base-pairs occluded per bound antibiotic molecule is calculated to be three, whereas for poly(dG)-poly(dC) it is estimated to be four to five. Poly(dA)-poly(dT) and poly(dI)-poly(dC) interact only very weakly with the antibiotic. Poly(dI-dC) interacts to a slightly greater extent, but the binding curve is quite unlike that seen with the three strongly binding synthetic polynucleotides. Echinomycin affects the supercoiling of closed circular duplex bacteriophage PM2 DNA in the characteristic fashion of intercalating drugs. At low ionic strength the unwinding angle is almost twice that of ethidium. Likewise the extension of the helix, determined from changes in the viscosity of rod-like sonicated DNA fragments, is nearly double that expected for a simple (monofunctional) intercalation process. On this basis the interaction process is characterized as bifunctional intercalation. At higher ionic strength the unwinding angle relative to that of ethidium and the helix extension per bound echinomycin molecule fall, indicating a smooth progression towards more nearly monofunctional intercalation. Two simpler compounds which act as analogues of the quinoxaline chromophores of echinomycin, quinoxaline-2-carboxamide and the trypanocidal drug Bayer 7602, interact with DNA very much more weakly than does echinomycin, showing that the peptide portion of the antibiotic plays an essential role in determining the strength and specificity of the interaction.

A map of the sites on bacteriophage PM2 DNA for the restriction endonucleases HindIII and HpaII

The map of the seven sites for the restriction endonuclease HindIII ‡ ‡ The nomenclature used for restriction endonucleases suggested by Smith & Nathans (1973) is endo R. HindIII, a restriction endonuclease from Hemophilus influenza; endo R. HpaII, a restriction endonuclease from H. parainfluenza. These have been further abbreviated to HindIII and HpaII in the interests of readability. and the single site for endo R. HpaII on PM2 DNA was determined. This map was oriented with respect to the denaturation map of this DNA (Brack et al., 1975) by partial denaturation mapping of the fragments. A new method for localizing restriction fragments by DNA-DNA hybridization and electron microscopy is described.

Characterization of DNA polymerase induced by bacteriophage T5 with DNA containing single strand breaks.

DNA polymerase induced by bacteriophage T5 was purified and characterized using mainly circular duplex DNA of bacteriophage PM2 with single strand breaks formed by DNase I action. A purification procedure is described which has consistently yielded DNA polymerase preparations with only one detectable protein band after polyacrylamide gel electrophoresis of either native protein in Tris-glyase preparations utilized both denatured DNA and nicked DNA as primer-templates, although at 37 degrees the activity with denatured DNA was much greater. Polymerase activities with both kinds of primer-templates were shown to be associated with one phage-induced protein. DNA synthesis with nicked DNA as primer-template increased with increasing numbers of single strand breaks. Essentially all such breaks were repairable by ligase. Alkaline sucrose gradient centrifugation showed that synthesis occurred with the strand which had a single strand break as a primer yielding DNA longer than one phage DNA unit length. Newly synthesized DNA was covalently linked to the primer strand. Thus the synthesis very likely occurred by strand displacement; this is supported by electron micrographs shown in the Appendix.

Conformational analysis of deoxyribonucleic acid from PM2 bacteriophage. The effect of size on supercoil shape

Laser light-scattering studies of bacteriophage PM2 DNA showed the molecule to have mol.wt. 5.9 X 10(6) and root-mean -square radius 125 nm at an ionic strength of 0.2 mol/litre. Computer-generated curves compatible with these data were compared with the experimental interference curve for several structural models of the molecules. The data fit best to an asymmetric four-armed planar molecule in which all four arms emerge from or close to the one area of the molecule. This contrasts with the smaller DNA molecules investigated, which have shown a three-armed molecule, whose symmetry varies with primary structure.

Binding of radiation-induced phenylalanine radicals to DNA: influence on the biological activity of the DNA and on its sensitivity to the induction of breaks by gamma-rays.

When an aqueous solution of double-stranded DNA of bacteriophage PM2 containing phenylalanine and saturated with N2O is irradiated with gamma-rays, radiation-induced phenylalanine radicals are bound covalently. Under the conditions used, about 25 phenylalanine molecules may be bound per lethal hit. For single-stranded PM2 DNA, most of the phenylalanine radicals bound are non-lethal. Evidence is presented that, in double-stranded DNA, an appreciable fraction of the single-strand breaks is induced by phenylalanine radicals. Radiation products of phenylalanine and the phenylalanine bound to the DNA decrease the sensitivity of the DNA to the induction of single-strand breaks. There are indications that the high efficiency of protection by radiation products of phenylalanine is due to their positive charge, which will result in a relatively high concentrations fo these compounds in the vicinity of the negatively-charged DNA molecules.

Attachment of nascent RNA molecules to superhelical DNA.

Comparisons are made of the attachment of nascent RNA molecules synthesized in vitro with Escherichia coli RNA polymerase on native bacteriophage PM2 DNA, a double-helical circular DNA with −110 superhelical twists, and on a derivative of this DNA with no superhelical twists. When RNA synthesis is terminated on either DNA by adding EDTA to chelate the Mg2+ ions, the nascent (incompletely polymerized) RNA chains remain bound to the DNA by bonds that involve fewer than 20 nucleotides of the RNA molecule. Differences between the two kinds of DNA are found when the complexes with nascent RNA are treated with sodium dedecyl sulfate or other agents that dissociate proteins (e.g. 3 m -CsCl; 5 m -urea; phenol). With such treatments, all RNA is released from the non-superhelical DNA, whereas only 40% of the RNA molecules are released from the superhelical DNA. The other 60% become bound even more tightly to that DNA, apparently by means of a hybrid helix that can involve up to 600 nucleotides of the bound RNA. It is suggested that the hybrid helix forms spontaneously on the superhelical DNA after denaturing the enzyme because the unwinding of the DNA helix needed to create the RNA-DNA helix relieves some of the strain that is known to exist in the superhelical DNA. Since the extensive hybrid helix does not exist before adding the detergent, the intact structure of the enzyme appears to be important to prevent its formation. Evidently RNA polymerase has a site that dissociates the polymerized portion of the RNA from the DNA as the enzyme moves along the template.

On the degree of unwinding of the DNA helix by ethidium: II. Studies by electron microscopy

When a negatively twisted covalently closed DNA is annealed with single-stranded fragments of the same DNA, under proper conditions a loop (or loops) may form by the disruption of a segment (or segments) of base pairs between the complementary strands of the covalently closed DNA, and the formation of base pairs between the strands of the covalently closed DNA and the single-stranded fragments. Since such a process involves essentially no net gain or loss of the number of base pairs, it is driven by the free energy favoring the reduction of the number of superhelical turns. If the fragments are sufficiently long or are present at a sufficiently high concentration during annealing, the most stable product between a covalently closed DNA and the DNA fragments (under conditions favoring the formation of double-stranded DNA) is a looped molecule devoid of superhelical turns. The size of the looped region or regions, which can be measured by electron microscopy, provides a way to determine the degree of superhelicity of the covalently closed DNA in the absence of the fragments. When this is compared with the degree of superhelicity of the covalently closed DNA determined by titration with the intercalative dye ethidium, the unwinding angle of the DNA double helix due to the intercalation of an ethidium can be calculated. Such measurements were done on two samples of phage PM2 DNA with different extents of supercoiling. The results are in agreement with the value 26° obtained recently by alkaline titration of covalently closed PM2 DNA samples in CsCl density gradients (Wang, J.C., (1974) J. Mol. Biol. 89, 783–801).

A freeze-squeeze method for recovering long DNA from agarose gels

Long DNA can be recovered from agarose gels after electrophoresis by freezing the gel slices and manually squeezing out liquid containing the DNA. With this method the recoveries of phage T 7 DNA (molecular weight 25 × 10 6) and the open and closed forms of circular phage PM2 DNA (molecular weight 6 × 10 6) were about 70%. Sedimentation analysis shows that the extruded DNA has not sustained double- or single-stranded breaks. The extruded DNA can be used without further purification as substrate for the restriction endonuclease Hind II,III, from Hemophilus influenzae, for DNA·DNA hybridization and for electron microscopy.

Initiation of transcription by Escherichia coli RNA polymerase from supercoiled and non-supercoiled bacteriophage PM2 DNA

Native bacteriophage PM2 DNA, a circular double-helical DNA with —50 superhelical twists, and derivatives of PM2 DNA containing no superhelical twists are compared for their ability to form stable binary complexes with Escherichia coli RNA polymerase. The number of enzyme molecules that can form such complexes, as measured by the number of chains initiated with ATP and GTP in the presence of saturating amounts of enzyme and using heparin to inactivate the unbound enzyme, is 16 per molecule of native PM2 DNA but only about two per molecule of closed non-supercoiled PM2 DNA. In each case equal numbers of chains are initiated with GTP and ATP. The reactivity of the sites on the two forms are also different; the rate of formation and the stability of the complexes are both lower on the non-supercoiled derivatives than on native PM2 DNA. The derivatives also contain a more significant proportion of sites that bind RNA polymerase tightly in an inactive state than native PM2 DNA. Thus, even with excess DNA (so that the number of sites is not limiting) and with sufficient preincubation to form the complexes, the non-supercoiled forms of DNA are still less active templates than native PM2 DNA. On the other hand, the active sites on the two DNAs are similar in one respect; the rates of RNA chain initiation by RNA polymerase bound to these sites are the same.

DNA-dependent RNA polymerase from Pseudomonas BAL-31. II. Transcription of the allomorphic forms of bacteriophage PM2 DNA.

Transcription of the supercoiled form (I) and the relaxed circular form (II) of bacteriophage PM2 DNA was studied utilizing the DNA-dependent RNA polymerase from its host, Pseudomonas BAL-31. Transcription of both templates is continuous for up to 2 hr, but proceeds at a two-fold higher rate on I than on II. This difference is mainly due to a 2.2-fold higher rate of chain initiation on I. When rifampicin (Rif) is added ater 10 min of synthesis, (1) transcription of II ceases by 30 min with a maximum product length of 7000 nucleotides (number average) being produced; (2) transcription of I continues with little rate reduction and with the product reaching 16,000 nucleotides (number average) by 2 hr. Sucrose gradient analysis shows that the product of II achieves maximum size 20 min after Rif addition and sediments in three peaks of 24, 33, and 39 S (approximately one-third, two-thirds, and one genome lengths). The product of I has a heterogeneous distribution and grows continuously with a large fraction reacting greater than 3 genome lengths by 90 min. The same differences in synthesis kinetics, Rif inhibition, and product size distribution are observed when I and II are transcribed by Escherichia coli RNA polymerase. These experiments show that (i) PM2 form I DNA is transcribed mainly by a process of continuous chain elongation, with little chain termination occurring; (ii) PM2 form II is transcribed by a process of continuous chain initiation, elongation, and termination of yield discrete products. Thus, the tertiary structure of circular DNA influences chain termination by RNA polymerase.

The degree of unwinding of the DNA helix by ethidium: I. Titration of twisted PM2 DNA molecules in alkaline cesium chloride density gradients

A series of covalently closed bacteriophage PM2 DNA samples with varying degrees of superhelicity were prepared in vitro. The amount of bound ethidium per DNA nueleotide needed for the removal of all superhelical turns, vc0, was determined for each sample by a number of methods. In order to evaluate the unwinding angle for the binding of one ethidium molecule to a DNA double helix, the pH dependence of the buoyant densities in CsCI of these samples was examined. A new calibration relating the change in buoyant density of a DNA to the fraction of bases titrated has been obtained, by measuring the buoyant densities of a number of catenanes (interlocked rings) containing both single-stranded and double-stranded λ DNA rings, at a pH such that the single-stranded DNA is fully titrated while the double-stranded DNA is not titrated. This calibration was used to obtain the pH dependence of the fraction of DNA bases titrated for the phage PM2 DNAs with differing extents of supercoiling. A simple theoretical analysis shows that in a restricted pH range close to pHm, the melting pH of the DNA in the absence of the topological constraint associated with covalently closed double-stranded DNAs, the difference in the fraction of bases titrated at a certain pH between two covalently closed DNAs with different degrees of superhelicity is directly proportional to the difference in the vc0 values of the DNAs. The unwinding angle per bound ethidium molecule can be obtained from the proportionality constant. In this way, it is not necessary to know precisely the actual pH value for either DNA, pHe, at which the DNA is titrated to the extent that it contains no superhelical turns. The conclusion of the theoretical analysis and the experimental results is that the binding of an ethidium molecule to a double-stranded DNA unwinds the DNA helix by an angle φe = 26 °. The uncertainty in this value is estimated to be less than 10%. The new value for φe is approximately a factor of two larger than the value 12 °, which has been in use in the past decade. In the earlier alkaline titration results for polyoma DNA (Vinograd et al., 1968), which had been interpreted as supporting the 12 ° value, the calculation of φe was critically dependent on knowing pHe. It is believed that pHe was underestimated in the earlier work, resulting in a low φe value. Since the previous value φe = 12 ° has been widely used in the determination of the number of superhelical turns for many DNAs, and in measurements on the angular alterations of the DNA helix by the binding of a variety of small and large molecules and by solvent and temperature changes, the new value φe = 26 ° requires proportional adjustments of many previous results.

Interactions between twisted DNAs and enzymes: The effects of superhelical turns

Covalently closed bacteriophage PM2 DNA samples containing varying numbers of superhelical turns were used as substrates for the holo and core enzymes of Escherichia coli RNA polymerase, E. coli DNA polymerase I, pancreatic DNAase, and two single strand-specific endonucleases from Neurospora crassa and Mung bean. The results indicate that the superhelical turns influence the properties of DNAs as enzyme substrates in two major ways: (1) if the interaction between a DNA and an enzyme involves the unwinding or winding of the DNA helix, the interaction is strongly affected by the degree of superhelicity of the DNA, since the higher free energy of a twisted DNA favors the reduction of superhelical turns. For phage PM2 DNAs with negative superhelical density, − σ 0 less than about 0.04, this factor is the dominant one. In this range of superhelical density the sensitivity of the DNAs toward endonucleolytic attack by the single strand-specific nucleates increases only slightly with increasing − σ 0. The magnitude of transcription by E. coli RNA polymerase holoenzyme, which is known to unwind the DNA helix slightly, increases rapidly with increasing − σ 0. A similar increase in template activity with increasing − σ 0 has been observed with the core enzyme, and is interpreted as evidence that the productive binding of the core polymerase also requires an unwinding of the DNA helix. (2) When − σ 0 is greater than about 0.04, it appears that the DNA is very tightly wound. Such tight twisting might reduce the binding and/or hinder the movement of certain macro-molecules along the helix. Thus, in this range of superhelix density, the template activity appears to decrease with increasing − σ 0. While the sensitivity of the DNA toward endonucleolytic scission by pancreatic DNAase is independent of σ 0, the sensitivity toward endonucleolytic scission by the single strand-specific nucleases increases markedly with increasing − σ 0 in this range. This is interpreted as resulting from large distortions of the double helix at the sharp bends at which the DNA doubles on itself because of the tight twisting. Such distorted regions become favored sites for the single strand-specific nucleases. For the DNA samples with − σ 0 as high as 0.06, there is no evidence that there is a significant fraction of bases in the single-stranded form. In the presence of oligo-nucleotides as initiators, no significant amount of DNA synthesis on twisted PM2 templates is detected with E. coli DNA polymerase I. While the highly twisted DNA with − σ 0 = 0.06 is nicked by the Mung bean enzyme about 75-fold faster than untwisted PM2 DNA, the rate is still only a few tenths of 1% of that with a single-stranded DNA as the substrate.

Ribonuclease-sensitivity of covalently closed rat liver mitochondrial deoxyribonucleic acid

Preparations of covalently closed mitochondrial DNA of rat liver contain 10-30% of molecules that are converted into relaxed circular molecules after treatment with ribonuclease. Control experiments, with covalently closed bacteriophage PM2 DNA, indicate that ribonuclease-sensitivity cannot be induced either by depurination or by incubation with reducing agents.

Dependence of the sedimentation coefficient of denatured closed circular DNA in alkali on the degree of strand interwinding. The absolute sense of supercoils

The sedimentation coefficients of closed circular Simian virus (SV40) DNA, phage PM2 DNA and animal mitochondrial DNAs in alkaline NaCl and alkaline CsCl were found to decrease by about 5% as the initial superhelix densities decreased from 0.0 to −0.10, corresponding to a decrease in the degree of strand interwinding from 1.0 to 0.9 net turns per ten base pairs. The small dependence of the appropriately normalized sedimentation coefficients on the degree of strand interwinding is taken to indicate that fully titrated and denatured closed circular DNA is highly supercoiled in a positive sense. This supercoiling results from the spontaneous decrease in the number of secondary turns in the no longer ordered pairs of polynucleotide strands. The measured sedimentation coefficients form a smoothly connected monotonie curve when plotted along with the sedimentation coefficients in alkali (Sebring et al., 1971) of parental closed circles derived from closed circular SV40 DNA replicating intermediates. These DNAs have degrees of strand interwinding that range from 0.6 to 0.15. The possibility raised by Paoletti & LePecq (1971) that closed circular duplex DNAs contain positive supercoils, i.e. have degrees of strand interwinding greater than 1.0, has been ruled out in a series of ethidium bromide titrations of partially replicated mitochondrial DNA before and after removal of the progeny strand. More ethidium bromide was required in the latter case for relaxation, a result which shows that intercalated ethidium and a displacing strand act on the duplex in the same way, and that both unwind the duplex. This result requires the supercoils of naturally closed circular DNAs to be negative.

Contribution of various types of damage to inactivation of bacteriophage PM2 DNA in frozen solution by gamma-radiation.

SummaryWhen biologically-active double-stranded circular DNA of the bacteriophage PM2 in a solution of nutrient broth is irradiated at −196°C by 60Co gamma-rays, about 85 per cent of the inactivation is due to nucleotide damage, i.e. damage in the nucleotides not associated with a chain break. Double-strand breaks contribute some 10 per cent to lethality and single-strand breaks only a minor fraction, if at all. About 10 per cent of the nucleotide damage that is lethal if present in single-stranded DNA is lethal in the double-stranded PM2 DNA. These results do not show large differences from those obtained previously for irradiation of PM2 DNA under conditions of indirect effect.

Further analysis of the altered secondary structure of superhelical DNA. Sensitivity to methylmercuric hydroxide a chemical probe for unpaired bases

A previous study in our laboratory of the reaction of formaldehyde with super-helical DNAs (φX replicative form and PM2) has led to a model for superhelical DNA in which there is a region or regions of altered secondary structure containing unpaired bases. Similar experiments using the nicked circular DNA gave no evidence of interruptions of base pairing. In this study we present additional data, which support the above model as well as extending our analysis of the secondary structure of superhelical DNA and the dynamics of the early denaturation process. In a series of experiments involving the binding of methyl-mercury as a chemical probe of unpaired bases, we obtained the following results. (1) Initially, both s 0 20w and the buoyant density of the superhelical form of phage PM2 DNA increased as a function of methylmercuric hydroxide concentration, whereas the nicked form did not. (2) This initial binding is accompanied by an increase in superhelical content τ from −41 to −46 turns. (3) The binding analysis allows us to estimate that 3.7% of the bases contain methylmercury in this phase of the transition. This is in excellent agreement with the extent of formylation. (4) Such a preformylated molecule shows a shift in the transition to lower mercurial concentrations. These results are interpreted as follows. The initial increase in −τ excludes the possibility that binding occurs to normal base-paired structures, since this would produce a coupled unwinding of duplex and superhelical turns. The additive effects of formylation and methylmercury binding support the concept that both chemical probes attack the same sites and induce similar structural changes. Thus the evidence clearly supports the view that superhelical DNA contains localized region(s) of interrupted base pairing. Recent studies from other laboratories using single strand-specific endonucleases are in complete agreement with this model.

Structure of DNA in denaturing solvents: I. Bacteriophage PM2 DNA in aqueous sodium perchlorate

The sedimentation velocity behavior of both closed (I) and nicked (II) circular bacteriophage PM2 DNA's has been determined in aqueous NaClO 4 solutions ranging from 0.98 to 7.2 m. The addition of the salt results in a progressive, non-cooperative unwinding of the DNA duplex to the extent of 3.9% at 7.2 m-NaClO 4, as determined by comparison with the sedimentation velocity/dye titration results of Upholt, Gray & Vinograd (1971), The salt-induced unwinding is complemented and extended by the intercalation of ethidium bromide, and the shape of the sedimentation velocity/dye titration curve in 0.98 m-NaClO 4 is similar to that in 2.85 m-CsCl ( Upholt et al., 1971) . The sedimentation coefficient ratio S I S II has the same dependence upon NaClO 4 concentration at 5 °C and at 20 °C, except for an effective translation to 0.8 m higher salt concentrations at the lower temperature. This shift is consistent with the known effects of temperature upon the duplex rotation angle ( Wang, 1969). The unwinding of the DNA duplex in the presence of increasing NaClO 4 concentrations is to be contrasted with the much smaller overwinding which occurs upon the addition of NaCl or CsCl at 20 °C ( Wang, 1969). The NaClO 4-induced unwinding takes place at temperatures well below the onset of the melting transition, and the specific salt effect does not appear to have a large temperature coefficient. The results indicate that substantial angular alteration of the duplex may be induced by a change in solution environment in the absence of transition to the coiled form of DNA.

Uncoiling of bacteriophage PM2 DNA by binding of steroidal diamines

Binding of irehdiamine A to closed circular duplex PM2 DNA results in removal and reversal of the supercoiling in a fashion qualitatively similar to that seen with intercalating drugs. The irehdiamine A-induced uncoiling has been measured in two buffers where the binding constants are known. Comparison with the effects of ethidium bromide in the same solvents yields estimates for the unwinding angle per bound irehdiamine A molecule of 5.1°±1.4° and 4.7°±1.5°. The uncoiling effect of irehdiamine A is associated with the formation of the primary (stabilising) complex since it also occurs with a bis-quaternised analogue, malouetine. The finding of a 5° unwinding angle effectively rules out any simple intercalation-type model for the interaction between irehdiamine A and DNA.

Infection of spheroplasts of Pseudomonas with DNA of bacteriophage PM2.

Spheroplasts of Pseudomonas BAL-31/PM2, obtained by treatment of the bacteria with lysozyme, can be infected with purified DNA from bacteriophage PM2. After 4 h of incubation the yield of progeny phage reaches a value of 107-6×107 plaque forming units/μg PM2 DNA. The yield increases linearly with the concentration of DNA over at least 3 orders of magnitude.

The superhelix density of bacteriophage PM2 DNA, determined by a viscometric method

FEBS LettersVolume 14, Issue 2 p. 125-129 Full-length articleFree Access The superhelix density of bacteriophage PM2 DNA, determined by a viscometric method E.M. Smit, E.M. Smit Department of Medical Enzymology, Laboratory of Biochemistry, University of Amsterdam, Amsterdam, The Netherlands 1Search for more papers by this authorP. Borst, P. Borst Department of Medical Enzymology, Laboratory of Biochemistry, University of Amsterdam, Amsterdam, The Netherlands 1Search for more papers by this author E.M. Smit, E.M. Smit Department of Medical Enzymology, Laboratory of Biochemistry, University of Amsterdam, Amsterdam, The Netherlands 1Search for more papers by this authorP. Borst, P. Borst Department of Medical Enzymology, Laboratory of Biochemistry, University of Amsterdam, Amsterdam, The Netherlands 1Search for more papers by this author First published: April 01, 1971 https://doi.org/10.1016/0014-5793(71)80117-5Citations: 17 Jan Swammerdam Institute, Eerste Constantijn Huygensstraat 20, Amsterdam, The Netherlands. AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL References 1 R. Weil, J. Vinograd, Proc. Natl. Acad. Sci. U.S., 50, (1963), 730– 2 P.H. Pouwels, C.M. Knynenburg, J. van Rotterdam, J.A. Cohen, H.S. Jansz, J. Mol. Biol., 32, (1967), 169– 3 L.V. Crawford, P.H. Black, Virology, 24, (1964), 388– 4 P. Borst, E.F.J. van Bruggen, G.J.C.M. Ruttenberg, A.M. Kroon, Biochim. Biophys. Acta, 149, (1967), 156– 5 L. Piko, A. Tyler, J. Vinograd, Biol. Bull., 132, (1967), 68– 6 D.S. Ray, A. Preuss, P.H. Hofschneider, J. Mol. Biol., 21, (1966), 485– 7 J.C. Wang, D. Baumgarten, B.M. Olivera, Proc. Natl. Acad. Sci. U.S., 58, (1967), 1852– 8 W. Bauer, J. Vinograd, J. Mol. Biol., 33, (1968), 141– 9 L.V. Crawford, M.J. Waring, J. Mol. Biol., 25, (1967), 23– 10 G.J.C.M. Ruttenberg, E.M. Smit, P. Borst, E.F.J. van Bruggen, Biochim. Biophys. Acta, 157, (1968), 429– 11 M.J. Waring, Nature, 219, (1968), 1320– 12 W. Bauer, J. Vinograd, J. Mol. Biol., 47, (1970), 419– 13 J.C. Wang, J. Mol. Biol., 43, (1969), 25– 14 A. Opschoor, P.H. Pouwels, C.M. Knynenburg, J.B.T. Aten, J. Mol. Biol., 37, (1968), 13– 15 R.T. Espejo, E.S. Canelo, Virology, 34, (1968), 738– 16 R.T. Espejo, E.S. Canelo, R.L. Sinsheimer, Proc. Natl. Acad. Sci. U.S., 63, (1969), 1164– 17 B.H. Zimm, D.M. Crothers, Proc. Natl. Acad. Sci. U.S., 48, (1961), 905– 18 J.B. Le Pecq, C. Paoletti, J. Mol. Biol., 27, (1967), 87– 19 M.J. Waring, J. Mol. Biol., 13, (1965), 269– 20 J.S. Wang, J. Mol. Biol., 43, (1969), 263– 21 H. Bujard, J. Mol. Biol., 33, (1968), 503– 22 E.M. Smit, unpublished experiments. 23 B. Hudson, W.B. Upholt, J. Devinny, J. Vinograd, Proc. Natl. Acad. Sci. U.S., 62, (1969), 813– 24 J. Vinograd, J. Lebowitz, R. Watson, J. Mol. Biol., 33, (1968), 173– 25 B.M.J. Révet, M. Schmir, J. Vinograd, Nature, 229, (1971), 10– Citing Literature Volume14, Issue2April 01, 1971Pages 125-129 ReferencesRelatedInformation