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. 1982 Jan 22;10(2):525–538. doi: 10.1093/nar/10.2.525

Determination of the number of superhelical turns by the hyperchromicity of partially denatured covalently-closed DNA molecules.

G Dougherty, T Koller
PMCID: PMC326155  PMID: 6895940

Abstract

A novel method of determining the number of superhelical turns of a covalently-closed plasmid DNA is described. It relies on the determination of the hyperchromicity, and hence the proportion of unstacked basepairs, of a partially heat-denatured sample which co-migrates during electrophoresis with nicked circular duplex DNA. The values obtained for plasmid pBR beta G DNA at 4 degrees C (-29.8 and -33.5 in the two buffers used) agree closely with the values obtained in parallel by topoisomer band-counting. Our method is less precise than band-counting but is readily applicable to determining the superhelicity of very large DNA molecules. Our results confirm earlier findings that magnesium-containing buffers cause an increase in the duplex winding angle, and hence an increase in the number of negative superhelical turns.

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Selected References

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