scholarly journals Non-intertwined strands of plasmid DNA contradict the Watson and Crick model of DNA structure

F1000Research ◽  
2020 ◽  
Vol 8 ◽  
pp. 356
Author(s):  
Pawan Kumar
Keyword(s):  
Electrophoretic Mobility ◽  
Plasmid Dna ◽  
Dna Structure ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
S1 Nuclease ◽  
Circular Dna ◽  
Exonuclease I ◽  
Alternative Structures ◽  
Native Plasmid

According to Watson and Crick (W/C) model of DNA structure, a DNA molecule consists of two antiparallel polynucleotide chains, intertwined with each other. Although W/C model is accepted widely, some researchers have raised questions against it and proposed alternative structures for DNA. In the present study, we examined W/C model using plasmid DNA. It was hypothesized that two strands of plasmid will remain intertwined (and not separate from each other) under denaturing conditions if it follows W/C model. To test this, plasmid DNA was denatured using sodium hydroxide (NaOH) and analyzed by agarose gel electrophoresis. It was observed that addition of 0.5 N NaOH to pUC19 or pBR322 plasmids resulted in a new form of DNA having higher electrophoretic mobility in agarose gel. Higher electrophoretic mobility DNA (HmDNA) in NaOH-denatured pUC19 was digestible with S1 nuclease, but not with HindIII and ‘exonuclease I + alkaline phosphatase’. These results demonstrated that HmDNA is single-stranded circular DNA, formed due to separation of two strands of NaOH-denatured plasmid. Single-stranded and circular nature of HmDNA was corroborated by its comparable electrophoretic mobility with purified top and bottom strands of plasmid DNA. Next, we examined whether HmDNA can re-anneal into the native plasmid. Interestingly, when subjected to renaturing conditions, HmDNA from NaOH-denatured pUC19 re-annealed to form native pUC19 plasmid, which was digestible with HindIII and induced ampicillin resistance in Escherichia coli. These findings demonstrated the reversible separation of two strands of plasmid DNA and contradicted the W/C model of DNA structure.

scholarly journals Non-intertwined strands of plasmid DNA contradicts the Watson and Crick model of DNA structure

F1000Research ◽  
2020 ◽  
Vol 8 ◽  
pp. 356
Author(s):  
Pawan Kumar
Keyword(s):  
Electrophoretic Mobility ◽  
Plasmid Dna ◽  
Dna Structure ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
S1 Nuclease ◽  
Circular Dna ◽  
Exonuclease I ◽  
Alternative Structures ◽  
Native Plasmid

According to Watson and Crick (W/C) model of DNA structure, a DNA molecule consists of two antiparallel polynucleotide chains, intertwined with each other. Although W/C model is accepted widely, some researchers have raised questions against it and proposed alternative structures for DNA. In the present study, we examined W/C model using plasmid DNA. It was hypothesized that two strands of plasmid will remain intertwined (and not separate from each other) under denaturing conditions if it follows W/C model. To test this, plasmid DNA was denatured using sodium hydroxide (NaOH) and analyzed by agarose gel electrophoresis. It was observed that addition of NaOH to pUC19 and pBR322 plasmids resulted in a new form of DNA having higher electrophoretic mobility in agarose gel. Higher electrophoretic mobility DNA (HmDNA) in NaOH-denatured pUC19 was digestible with S1 nuclease, but not with HindIII and ‘exonuclease I + alkaline phosphatase’. These results demonstrated that HmDNA is single-stranded circular DNA, formed due to separation of two strands of NaOH-denatured plasmid. Single-stranded and circular nature of HmDNA was corroborated by its comparable electrophoretic mobility with purified top and bottom strands of plasmid DNA. Next, we examined whether HmDNA can re-anneal into the native plasmid. Interestingly, when HmDNA from NaOH-denatured pUC19 was subjected to renaturing conditions, it formed native pUC19 plasmid, which was digestible with HindIII and induced ampicillin resistance in Escherichia coli cells. These findings demonstrated the reversible separation of two strands of plasmid DNA and contradicted the W/C model of DNA structure.

scholarly journals Non-intertwined strands of plasmid DNA contradicts the Watson and Crick model of DNA structure

F1000Research ◽  
2019 ◽  
Vol 8 ◽  
pp. 356
Author(s):  
Pawan Kumar
Keyword(s):  
Electrophoretic Mobility ◽  
Plasmid Dna ◽  
Gel Electrophoresis ◽  
Dna Structure ◽  
Agarose Gel ◽  
Circular Dna ◽  
Alternative Structures ◽  
Dna Molecule ◽  
Dna Strands ◽  
New Form

According to Watson and Crick (W/C) model, a DNA molecule consists of two antiparallel polynucleotide chains, intertwined with each other. Although the W/C model is accepted widely, a number of researchers have raised questions against it and proposed alternative structures for DNA. In the present study, the W/C model was examined using plasmid DNA. It was hypothesized that two strands of plasmid DNA will remain intertwined and not separate from each other under denaturing conditions, if it follows the W/C model. To test this, plasmid DNA was denatured using sodium hydroxide (NaOH) and analyzed by gel electrophoresis. It was observed that addition of NaOH to pUC19 and pBR322 plasmids resulted in new form of DNA with higher electrophoretic mobility in agarose gel. DNA corresponding to higher electrophoretic mobility band of pUC19 (hmP19) was single-stranded and circular, indicating the separation of two strands of pUC19 plasmid. Next, we examined whether hmP19 DNA can re-anneal to form native pUC19 plasmid. It was observed that neutralization of NaOH resulted in the appearance of native pUC19 plasmid in denatured DNA. Native pUC19 was also formed by hmP19 DNA extracted from agarose gel and was found to be digestible with Hind III. Ability to confer ampicillin resistance in transformed Escherichia coli demonstrated the functionality of pUC19 plasmid formed by extracted hmP19 DNA. Reversible separation of two strands of plasmid into single-stranded circular DNA shows that DNA strands are not intertwined with each other and contradicts the W/C model of DNA structure.

Simple procedure for distinguishing CCC, OC, and L forms of plasmid DNA by agarose gel electrophoresis

Plasmid ◽  
1981 ◽  
Vol 5 (3) ◽  
pp. 371-373 ◽  
Author(s):  
G. Hintermann ◽  
H.-M. Fischer ◽  
R. Crameri ◽  
R. Hütter
Keyword(s):  
Plasmid Dna ◽  
Gel Electrophoresis ◽  
Simple Procedure ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis

Plasmid detection in a bacteriocinogenic strain of Clostridium perfringens

1980 ◽  
Vol 26 (8) ◽  
pp. 1018-1022 ◽  
Author(s):  
A. W. Li ◽  
P. J. Krell ◽  
D. E. Mahony
Keyword(s):  
Clostridium Perfringens ◽  
Plasmid Dna ◽  
Gel Electrophoresis ◽  
Density Gradient Centrifugation ◽  
Gradient Centrifugation ◽  
Buoyant Density ◽  
Size Distributions ◽  
Agarose Gel Electrophoresis ◽  
Circular Dna ◽  
Molecular Weights

Bacteriocinogenic Clostridium perfringens, strain 28, harboured plasmid DNA detectable by dye-buoyant density-gradient centrifugation. This plasmid DNA was absent from an ultraviolet light cured variant which had simultaneously lost its immunity and ability to produce bacteriocin. Agarose gel electrophoresis of the plasmid DNA revealed at least six bands but denaturation experiments suggested three plasmids occurring in more than one conformation. Electron microscopy revealed three major size distributions of circular DNA of molecular weights 1,5.6, and 7.1 megadaltons. Some evidence suggests that the 5.6 megadalton plasmid may control bacteriocin 28 production.

scholarly journals Plasmid DNA replication and topology as visualized by two-dimensional agarose gel electrophoresis

Plasmid ◽  
2010 ◽  
Vol 63 (1) ◽  
pp. 1-10 ◽  
Author(s):  
J.B. Schvartzman ◽  
M.L. Martínez-Robles ◽  
P. Hernández ◽  
D.B. Krimer
Keyword(s):  
Dna Replication ◽  
Plasmid Dna ◽  
Gel Electrophoresis ◽  
Agarose Gel ◽  
Two Dimensional ◽  
Agarose Gel Electrophoresis ◽  
And Topology

Agarose gel electrophoresis of circular DNA of replicative form of bacteriophage G4

1977 ◽  
Vol 78 (1) ◽  
pp. 260-275 ◽  
Author(s):  
Frances C. Wheeler ◽  
Richard A. Fishel ◽  
Robert C. Warner
Keyword(s):  
Gel Electrophoresis ◽  
Agarose Gel ◽  
Agarose Gel Electrophoresis ◽  
Replicative Form ◽  
Circular Dna

Characterization and transformation of plasmid pAA-1 found in an antarctic cryptoendolithic bacterium

1996 ◽  
Vol 42 (6) ◽  
pp. 571-576 ◽  
Author(s):  
Christian G. Gliesche ◽  
Marina Jendrach ◽  
Klaus Peissl ◽  
Jörg Siebert ◽  
Peter Hirsch
Keyword(s):  
Plasmid Dna ◽  
Gel Electrophoresis ◽  
Natural Transformation ◽  
Restriction Map ◽  
Agarose Gel ◽  
Dry Valleys ◽  
Agarose Gel Electrophoresis ◽  
Bacterial Isolates ◽  
E Coli ◽  
Natural Genetic Transformation

Sixty-three bacterial isolates from antarctic sandstone samples (Linnaeus Terrace, Asgard Range, McMurdo Dry Valleys) were screened for the presence of plasmid DNA. Twenty-seven percent of all the isolates (mainly Gram-positives) harbored one or more plasmids of low molecular mass (1.1–16.3 kb). Strain AA-341 contained plasmid pAA-1 (2.9 kb), as demonstrated by agarose gel electrophoresis and restriction endonuclease digests. This plasmid conferred resistance to chromium and ampicillin. It was not conjugative, but it could be transferred by electroporation to chromium- and ampicillin-sensitive strains AA-330, AA-338, and E. coli HB101. A restriction map of pAA-1 was constructed with HindIII, SalI, ScaI, AvaII, EcoRI, PvuII, BamHI, and DraI. Electrotransfer of this plasmid from E. coli HB101(pAA-1) to strain AA-330 was demonstrated. By natural genetic transformation, plasmid pAA-1 could be transferred into the sensitive strain AA-334, which thereby became resistant to chromium and ampicillin. The importance of such processes for the colonization of stressed environments is discussed.Key words: Antarctica, cryptoendolithic bacteria, plasmids, resistance to chromium, natural transformation.

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